WO2008145875A1 - Fluorescent organic nanocrystals for producing biosensors - Google Patents

Fluorescent organic nanocrystals for producing biosensors Download PDF

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Publication number
WO2008145875A1
WO2008145875A1 PCT/FR2008/050490 FR2008050490W WO2008145875A1 WO 2008145875 A1 WO2008145875 A1 WO 2008145875A1 FR 2008050490 W FR2008050490 W FR 2008050490W WO 2008145875 A1 WO2008145875 A1 WO 2008145875A1
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Prior art keywords
nano
inorganic
organo
layer
organic
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PCT/FR2008/050490
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French (fr)
Inventor
Alain Ibanez
Virginie Monnier
Nathalie Sanz
Robert Pansu
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Centre National De La Recherche Scientifique (Cnrs)
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Priority to JP2009554073A priority Critical patent/JP5250569B2/en
Priority to US12/532,128 priority patent/US8367002B2/en
Priority to EP08805659A priority patent/EP2125992B1/en
Priority to CA2681300A priority patent/CA2681300C/en
Publication of WO2008145875A1 publication Critical patent/WO2008145875A1/en

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    • CCHEMISTRY; METALLURGY
    • C09DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09KMATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K11/00Luminescent, e.g. electroluminescent, chemiluminescent materials
    • C09K11/02Use of particular materials as binders, particle coatings or suspension media therefor
    • C09K11/025Use of particular materials as binders, particle coatings or suspension media therefor non-luminescent particle coatings or suspension media
    • BPERFORMING OPERATIONS; TRANSPORTING
    • B82NANOTECHNOLOGY
    • B82YSPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y15/00Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
    • G01N33/585Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with a particulate label, e.g. coloured latex
    • G01N33/587Nanoparticles
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
    • G01N33/588Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with semiconductor nanocrystal label, e.g. quantum dots
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10TTECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T428/00Stock material or miscellaneous articles
    • Y10T428/24Structurally defined web or sheet [e.g., overall dimension, etc.]
    • Y10T428/24355Continuous and nonuniform or irregular surface on layer or component [e.g., roofing, etc.]

Definitions

  • the present invention relates to the field of materials with optical properties, and in particular sensors and sensors, including biological.
  • it may be chips, especially biochips, in particular fluorescence.
  • Biochips are major tools in clinical research, in particular for the development of diagnostic tests and prognoses, as well as in the search for new therapeutic procedures.
  • Biochips are fluorescence biochips. However, these may have insufficient sensitivity to analyze precious and expensive samples, fragile and / or complicated to manufacture. Biochips may still be insufficiently transparent, especially in certain wavelengths, such as Infra-Red. They may also be insufficiently stable, for example not have satisfactory chemical or temporal resistance.
  • the subject of the invention is a (nano) material comprising at least one inorganic and / or organo-mineral layer, in which at least one type of nanoparticles is integrated.
  • luminescent crystal at least a portion is in direct contact with the external environment.
  • the organic nano-crystals or organo-metallic luminescent are fluorescent nano-crystals.
  • the (nano) material comprises at least one inorganic and / or organo-mineral layer, in which is integrated at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of the said layer so that at least a portion of the nano-crystal is in direct contact with the external medium.
  • (nano) material is meant in the sense of the present invention a material or a nanomaterial as defined below.
  • the term “nanomaterial” means a material of nanometric size, that is to say a material of which at least one of the dimensions is at the nanoscale. In other words, it is a material whose nanometric size in at least one of the dimensions of the space.
  • the size of the material in at least one of the dimensions of the space is between 1 and 500 nm, preferably between 1 and 100 nm.
  • it may be a thin layer.
  • the term “organo-mineral layer” means a matrix comprising at least one mineral compound and at least one organic compound.
  • the organomineral layer may be a sol-gel layer, for example of the Silicate, Titanate, Zirconate, Stannate, Borate, Aluminate or Yttriate variety, and comprising at least organic or organometallic nanocrystals.
  • the sol-gel layer may in particular consist mainly of silicate / amorphous silicon oxide, which may result from the hydrolysis and polycondensation of the silicon alkoxides included in the initial solution.
  • sol-gel layer may also contain alkyl, aryl and / or aralkyl carbon radicals.
  • the inorganic and / or organo-mineral layer may especially comprise at least one sol-gel layer.
  • This sol-gel layer may especially be one of silicate or metal variety, in particular accessible from precursor alkoxides of silicon, titanium alkoxides or other metals, in particular selected from titanium, zirconium, tin, Boron, Aluminum and Yttrium.
  • the different types of alkoxides can be used at 100% or as a mixture in varying proportions.
  • the inorganic and / or organo-mineral layer comprising a sol-gel layer may be obtained from a sol-gel mixture.
  • the sol-gel mixture contains at least one fluorescent organic or organometallic compound.
  • this fluorescent organic or organometallic compound initially dissolved in the sol-gel mixture is crystallized in the sol-gel matrix to form nano-crystals.
  • organic or organo-metallic fluorescent The crystallization can be carried out by drying the sol-gel mixture containing the organic or organo-metallic fluorescent compound. The speed of drying can be adjusted to promote the formation of nano-crystals.
  • sol-gel is understood to mean a mixture initially comprising at least one metal alkoxide in the presence of water, at least one organic solvent and at least one fluorescent organic or organo-metallic compound, said mixture being initially in the form of a solution and producing a gel by hydrolysis and polycondensation reaction.
  • the presence of water makes it possible to initiate hydrolysis and condensation reactions of metal alkoxides forming an inorganic network, leading to gels and then xerogels. These xerogels lead to a structured inorganic solid network of metal oxides.
  • Sol-gel chemistry is for example cited in French patent FR 2,853,307 [ref 4].
  • the sol-gel layer can be obtained from any metal alkoxide known to those skilled in the art compatible with sol-gel chemistry.
  • the sol-gel layer can be obtained from a metal alkoxide of formula (I):
  • M is a metal selected from the group consisting of Si, Ti, Zr, Sn, B, Al and Y, for example Si x is an integer from 0 to 2, for example 0 or 1; y is an integer ranging from 1 to 6, for example 3 or 4; x + y corresponds to the coordination number of the metal M;
  • R 1 and R 2 independently represent an organic radical compatible with sol-gel chemistry.
  • R 1 and R 2 may independently represent an alkyl, aryl or aralkyl carbon radical.
  • the alkyl radicals may be linear or branched, cyclic or acyclic, saturated or unsaturated, and optionally include or carry heteroatoms, especially O, N, S, B and P. It may for example be C ⁇ alkyl; C 2 _ 15 alkenyl; C 2 _ 15 alkynyl; C 3 -C 15 cycloalkyl.
  • the alkyl radicals can comprise from 1 to 15 carbon atoms, and in particular be chosen from methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl and dodecyl radicals.
  • the alkyl radical can comprise 1 to 15 carbon atoms, for example 1 to 10 carbon atoms, for example 1 to 6 carbon atoms.
  • the alkyl radicals may comprise or carry one or more heteroatoms, in particular chosen from the group comprising oxygen, sulfur, nitrogen, phosphorus and boron.
  • the compounds of formula (I) in which R 1 and / or R 2 is an alkyl radical include the compounds of formula (I) in which R 1 and / or R 2 is a heteroalkyl radical. It may for example be a heteroalkyl, a heteroalkenyl, a heteroalkynyl, a heterocycle, an alkoxy, an alkylthio, an alkylamine, an alkylamide, a alkylimine, alkylimide, alkyl ester, alkyl ether, etc.
  • the heteroalkyl radical may comprise 1 to 15 carbon atoms, for example 1 to 10 carbon atoms, for example 1 to 6 carbon atoms.
  • aryl means a radical comprising at least one ring satisfying Hekel's aromaticity rule. Said aryl may be mono- or polycyclic, fused or not.
  • the aryl radicals may comprise from 4 to 15 carbon atoms, in particular from 6 to 10 carbon atoms, and optionally include heteroatoms, in particular O, N, and S.
  • the aryl radicals may comprise or carry one or more heteroatoms, in particular chosen from the group comprising oxygen, sulfur, nitrogen and phosphorus.
  • the compounds of formula (I) wherein R 1 and / or R 2 and an aryl radical include the compounds of formula (I) wherein R 1 and / or R 2 is a heteroaryl radical, that is to say an aromatic hydrocarbon ring or polycyclic group in which at least one carbon atom is replaced by a heteroatom, in particular chosen from the group comprising oxygen, sulfur and nitrogen.
  • aralkyl means an alkyl radical substituted with a radical. aryl, the terms “alkyl” and “aryl” being as defined above.
  • the point of attachment of the "aralkyl” radical to the remainder of the molecule i.e., to the metal M in formula (I)) is at the alkyl group.
  • the aralkyl radicals can comprise from 4 to 30 carbon atoms, in particular from 6 to 20 carbon atoms, and optionally comprise or be substituted by alkyl radicals and / or heteroatoms, in particular O, N, S and P.
  • Aralkyl radicals include benzyl and tolyl.
  • alkyl, aryl and / or aralkyl radicals may be bonded directly to the atoms of silicon, titanium, zirconium, tin, boron, aluminum, yttrium, in particular by a covalent bond.
  • hybrid organic-mineral materials as well as by examples are known materials in the field of contact lenses obtained by sol-gel.
  • the sol-gel layer may comprise only silicon, or silicon and titanium, zirconium and / or tin.
  • This sol-gel layer can in particular be obtained by using other types of metal alkoxides, such as titanium alkoxides, zirconium and / or
  • the metal alkoxides react via hydrolysis and condensation reactions, leading to "inorganic polymers”.
  • organic polymer In the field of sol-gel chemistry, the term "inorganic polymer” is used in analogy with organic polymers. By way of illustration, the following reactions of hydrolysis (1) and condensation (2) of metal alkoxides can take place.
  • the sol-gel mixture of the process of the invention can be homogeneous and stable chemically and mechanically. For example, it can be stable for several months or even years, which presents a great advantage for the practical implementation of the present invention.
  • metal alkoxides also known as "sol-gel precursors" within the meaning of the present invention, any metal compound of formula (I).
  • any metal alkoxide known to those skilled in the art can be used insofar as an interconnected polymeric sol-gel network can be obtained.
  • Those skilled in the art may in particular use any metal alkoxide specific to known sol-gel chemistry.
  • the reader may in particular refer to the reference book CJ. Brinker and GW Scherer, Sol-Gel Science, The Physics and Chemistry of SoIngI Processing, Academy Press, New York, 1990 [ref I]. See also ref 2 and 3.
  • solvent means any solvent known to those skilled in the art compatible with the process for implementing the present invention, that is to say to say that can be miscible metal alkoxides and hydrolysis water to obtain homogeneous solutions, especially suitable for their shaping thin film or film.
  • the solvent makes it possible to completely dissolve the fluorescent compound.
  • the water present in the sol-gel mixture can be introduced initially in a small quantity, this being sufficient to initiate the reactions leading to the polycondensation of the inorganic network, or in a quantity greater than the quantity initial metal alkoxide.
  • the sol-gel mixture may initially comprise a molar percentage of water relative to the number of alkoxide functional groups (-OR 2 ) of from 10 to 400%, preferably from 20 to 100%, more preferably from 50 to 100 %.
  • Silicate precursor alkoxides may be chosen, for example, from the group comprising tetramethoxysilane (TMOS, Si (OCH 3 ) 4 ), tetraethoxysilane (TEOS, Si (OC 2 H 5 ) 4 ), methyltrimethoxysilane (MTMOS, CH 3 Si (OCH 3 ) 3 ), methyltriethoxysilane (MTEOS, CH 3 Si (OC 2 H 5 ) 3 ), ethyltriethoxysilane (ETEOS, C 2 H 5 Si (OC 2 H 5 ) 3 ) 1,2-bis (trimethoxysilyl) ethane (TMSE), 3-glycidoxypropyl) trimethoxysilane (GPTMS), or a mixture thereof.
  • TMOS tetramethoxysilane
  • TEOS tetraethoxysilane
  • TEOS tetraethoxysilane
  • the non-linking methyl substituents of MTMOS can cover, at least in part, the pores of the gel and promote the aggregation of the nano-crystals, while the silanol groups can form bonds, in particular hydrogen bonds, with the organic molecules.
  • the inorganic and / or organo-mineral layer in particular the sol-gel layer, is in the form of a film.
  • film can be understood in the sense of the present invention a structure in which the thickness is of the order of a few hundred nanometers or the order of a micrometer, and in particular a thickness of 50 nm at 1500 nm, in particular from 80 to 1200 nm, or even from 100 to 1000 nm.
  • this film has a width and / or length of 1 mm to 10 cm, in particular 5 mm to 5 cm.
  • the nano-crystals may be fluorescent, especially they absorb and re-emit light in the wavelengths of the visible or near infrared.
  • the luminescence may spread over a set of luminescent molecules, or chromophores, for example ranging from 10 3 to 10 10 , in particular from 10 3 to 10 9 , in particular from 10 4 to 10 8 , or even from 10 5 to 10 9. 10 7 molecules per nano-crystal.
  • the nano-crystals emit a fluorescence, which is dispersed over all fluorophores and is quenched or quenched simultaneously.
  • fluorescence is dispersed over all fluorophores and is quenched or quenched simultaneously.
  • simultaneous is meant from 0.001 to 10 ns, in particular from 0.01 to 5 ns, in particular from 0.1 to 2 ns, or even from 0.5 to 1.5 ns.
  • the term "fluorescent" compound or nanocrystal means a compound or nanocrystal having the property of emitting, following a light excitation, electromagnetic radiation in the visible light domain or in the Near infrared (IR) -
  • the excitation light signal can come from a light source such as a laser, an arc lamp or light-emitting diodes (LEDs).
  • the fluorescent compound or nanocrystal can emit electromagnetic radiation at wavelengths of 400 to 1200 nm, which corresponds to the window of relative transparency of living tissues.
  • the fluorescent compound or nanocrystal may emit electromagnetic radiation at wavelengths of 400 to 1000 nm, for example from 550 to 800 nm, which corresponds to fluorescence in the red and the near infra-red.
  • the fluorescent compound or nanocrystal can emit fluorescence in the visible and near infra-red.
  • the nano-crystals may be organic or organometallic nano-crystals, in particular be fluorescent, in particular comprise at least one, or even consist of a fluorescent compound in the crystalline state.
  • the nano-crystals may comprise at least one organic molecule compound or coordination complex which fluoresces in the solid state and is soluble in an organic solvent.
  • the fluorescent organic or organo-metallic compound may for example be chosen from the group comprising the fluorescent compounds of the polyaromatic family, for example diphenylanthracene, rubrene or tetracene, the aromatic family, for example 4- (N, N-diethylamino) -b-nitrostyrene (DEANST), 1,4-di (2,5-phenyloxazole) benzene (POPOP) or N-4-nitrophenyl- (L) -prolinol ( NPP), the stilbene family, for example cyano-methoxy-nitrosilbene (CMONS), diethyl-amino-nitro-stilbene or stilbene, the family of naphthimides, for example naphthimide, the family of rhodamines, for example Rhodamine B, the family of auramines, for example auramine O, the family of perylene diimides, derivatives of dipyrromethene difluoride boron
  • the fluorescent compound in the solid state, or even crystalline may in particular be selected from the group comprising cyanomethoxynitrostilbene, naphthimide, polyaromatic molecules, such as rubrene or tetracene, perylenes diimides, diphenylanthracene, derivatives of dipyrromethene boron difluoride (Dipyrromethene Boron Difluoride - Bodipy®), and Europium complexes.
  • the fluorescent compound is selected from the polyaromatic family, such as rubrene, auramine O or rhodamine B, for example.
  • the nano-crystals can be inserted into the pores of the inorganic and / or organo-mineral layer.
  • pores is meant within the meaning of the present invention interstices or voids between the chains of the network forming the inorganic and / or organo-mineral layer (for example, the silicate network, titanate, zirconate, stannate, borate, aluminate, ytirate, etc.) which polymerized / polycondensed during the sol-gel process.
  • the silicate network titanate, zirconate, stannate, borate, aluminate, ytirate, etc.
  • the nano-crystals observed by transmission electron microscopy may be spherical in shape and may have a diameter ranging from 10 to 500 nm, in particular from 100 to 400 nm, in particular from 150 to 300 nm.
  • the nano-crystals may have a relatively homogeneous size distribution, in particular varying at most by +/- 20%, or even +/- 10%.
  • the nano-crystals present in the inorganic and / or organo-mineral layer have at least a part in direct contact with the outside, in particular they emerge from 2 to 50% of their diameter relative to the inorganic layer. and / or organo-mineral, or even 5 to 30% of their diameter, in particular when the nanocrystals are spherical.
  • the inorganic and / or organo-mineral layer may be a sol-gel matrix.
  • the nanocrystals may exceed the inorganic and / or organo-mineral layer from 1 to 100 nm, in particular from 2 to 80 nm, or even from 3 to 50 nm. It suffices that the nanocrystals emerge a little surface to be in direct contact with the outside (organic solution) is a few nm to a few tens of nm.
  • the emergence height may depend in part on the diameter of the nanocrystals.
  • the emergence height does not exceed one third of the diameter of the nanocrystals, especially so that they remain well anchored in the sol-gel layer. This measurement may in particular be performed by atomic force microscopy in intermittent oscillating mode by recording the topography of the sample.
  • the nano-crystals present in the inorganic and / or organo-mineral layer are firmly anchored in it, in particular they can withstand immersion in a biological solution to be analyzed (DNA strands, proteins. ..) and this for a few minutes.
  • resist means that more than 95%, especially more than 99% or more than 99.9% of the nanocrystals present at the beginning of the test are still present in the (nano) material at the end of the test.
  • these nano-crystals may have at their surface at least one type of probe.
  • Nano-crystals can have one or several types of identical or different probes, for example one, two, three, four or five identical or different probes, in particular one, two, three, four or five probes of different types.
  • At least one probe may especially be chosen from the group comprising color indicators, DNA, oligonucleotides or polynucleotides, polypeptides, proteins, antibodies, sugars, glycoproteins and lipids.
  • the colored indicators can be selected from pH, redox and / or complexation indicators.
  • This probe can be simply adsorbed or grafted onto the nano-crystals, in particular via a chemical bond.
  • the probe can be absorbed or grafted via in particular a covalent, ionic, electrostatic, ionocovalent bond, a hydrogen bond, hydrophobic interactions, van der waals forces.
  • the probe can be chemically modified with a suitable chemical function so as to be grafted on the surface of the nano-crystal.
  • the optionally chemically modified probe does not bind to the surface of the sol-gel layer, in particular it is insensitive to free M-OH sites present on the surface of the sol-gel layer.
  • emerging nano-crystals constituting, thanks to their fluorescence, the signaling function can thus be functionalized by grafting a biomolecule carrying a probe function, such half-strand of DNA, to thereby achieve a bio chip, sensor or sensor comprising a (nano) material according to the invention, or a support for bio-chip, sensor or sensor. This aspect of the invention is discussed in more detail in the sections that follow.
  • the subject of the invention is a process for the manufacture of (nano) material according to the invention, in particular comprising an inorganic and / or organo-mineral layer, in which at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of said layer so that at least a portion is in direct contact with the external medium
  • said inorganic and / or organo-mineral layer comprises or includes at least one type of fluorescent nano-crystal.
  • This step of partial dissolution of the sol-gel layer can be carried out by agents, in particular chemical agents, for homogeneously etching the surface of the inorganic and / or organo-mineral layer, in particular without deteriorating the properties of the fluorescence of nano-crystals.
  • agents for pickling selected from basic aqueous solutions, in particular inorganic or organic bases, strong or weak, including soda, ammonia and / or potassium hydroxide.
  • these basic solutions have a base concentration ranging from 10 "6-10" 1 million, including 10 "3-10" 1 M.
  • this step may allow a regular thinning of the inorganic and / or organo-mineral layer, by stripping the surface of the layer.
  • inorganic and / or organo-mineral, and in particular silicate sol-gel matrices may be stripping the surface of the layer.
  • this controlled dissolution step allows a thinning rate ranging from 5 to 100 nm / h, or 10 to 50 nm / h.
  • the method according to the invention may also comprise a step of preparing at least one matrix layer, in particular of sol-gel matrix, comprising or including nano-crystals.
  • the invention relates to a method of manufacturing (nano) material comprising an inorganic and / or organo-mineral layer, in which is integrated at least one type of nano-crystal organic or organometallic fluorescent emerging from the surface of said layer so that at least a portion is in direct contact with the external medium, said method comprising: a) a step of preparing at least one inorganic and / or organic layer -mineral comprising or including organic or organometallic fluorescent nano-crystals, and b) a step of dissolving a part of said inorganic and / or organo-mineral layer obtained in step a) so as to emerge at least part of the nano-crystals of the inorganic and / or organo-mineral layer or to increase the surface of the nano-crystals in contact with the external medium.
  • step a) may comprise a preliminary step (0) for preparing a sol-gel mixture comprising: (i) a step (Oa) for preparing an initial sol-gel mixture, for example by mixing in a solvent at least one fluorescent organic or organometallic compound and at least one metal alkoxide, for example of formula (I) in the presence of water; and eventually ,
  • the duration d may be less than several months or even several years.
  • the duration d can be from 1 day to 1 year, for example from 7 days to 21 days.
  • the storage of the mixture allows the mixture to age to properly advance the hydrolysis and condensation reactions of metal alkoxides.
  • the step (0) for preparing the sol-gel mixture may comprise the addition of an acid to the initial mixture.
  • the acid by lowering the pH of the mixture, makes it possible to promote the production of long inorganic chains which are favorable for the formation of a dense inorganic and / or organo-mineral layer around the organic or organometallic crystals.
  • the pH of the mixture can be from 1 to 7, preferably from 1 to 2.
  • the term "acid” means the Brönsted acids, inorganic or organic.
  • acids that can be used, mention may be made, for example, of hydrochloric acid, nitric acid or acetic acid.
  • the fluorescent organic or organo-metallic compound, the metal alkoxide and the solvent may be selected according to any one of the embodiments described above for the implementation of the nanomaterials of the invention.
  • Step a) may in particular be carried out by crystallization of organic nano-crystals or organometallic fluorescents in thin layers of sol-gel matrix.
  • this step can be carried out by "spin-coating" centrifugation.
  • the preparation of this inorganic and / or organo-mineral layer, optionally in the form of a thin layer can be carried out at room temperature by hydrolysis and condensation of a solution comprising precursors of the inorganic layer and / or or organo-mineral, in particular sol-gel, such as silicon alkoxides, in which the organic or organometallic molecules are dissolved in a solvent, in particular an organic solvent.
  • organo-metallic is meant compounds comprising an organic part and a metal part.
  • the nano-crystallization can result from a strong instantaneous nucleation, followed by a control of the growth of nuclei by the viscosity and / or the porosity of the gel.
  • the supersaturation of the organic phase can be caused by the very rapid evaporation of the organic solvent.
  • the viscosity of the solvent and or the deposition time of the layers in particular in the spin spin coating procedure, depending on the rotational speed, for example from 1000 to 4000 rpm, the viscosity of the solvent and or the deposition time of the layers.
  • Thin and / or transparent, for example 100 to 1000 nm can be made.
  • the inorganic and / or organo-mineral layer of step a) can then undergo an annealing step, especially at temperatures close to the melting point of the organic nano-crystals.
  • This annealing step is intended to eliminate any trace of solvent and further stabilize the sol-gel layer, in particular inorganic (residual solvent removal, densification of the matrix).
  • the annealing step may be carried out especially at temperatures of from 50 to 250 0 C, in particular from 80 to 150 0 C.
  • the term "temperature of the melting point” is meant within the meaning of the present invention a temperature ranging from a melting point of less than 5 ° C. to a melting point of less than 100 ° C., in particular of melting point less than 10 ° C. with a melting point of less than 50 ° C.
  • This annealing step may also make it possible to improve the crystallinity of the aggregates and / or the stabilization of the inorganic and / or organo-mineral layer.
  • the size of the nano-crystals can thus be adjusted according to the conditions of nano-crystallization confined in the inorganic and / or organo-mineral layer, in particular in the sol-gel matrix, in particular silicate.
  • nano-crystals are fluorescent, they can be visualized by techniques such as confocal photonic microscopy. This is illustrated in FIG. 2 which is an image by confocal photon microscopy of nano-rubrene crystals in sol-gel matrix.
  • chromophores organic or organo-metallic fluorescent molecules
  • sol-gel matrix for example the nature of the silicon alkoxides, the kinetics of hydrolysis and condensation, the nature and levels of solvents, the concentration of fluorophores, the supersaturation and the centrifugation parameters.
  • the present invention relates to a film of (nano) material according to the invention.
  • it is a film comprising a sol-gel matrix comprising mechanically stabilized nano-crystals, said nano-crystals emerging from the surface of said film so that at least a portion of each nano -crystal is in direct contact with the external environment.
  • the (nano) material is a sol-gel matrix comprising mechanically stabilized nano-crystals, said nano-crystals emerging from the surface of said film so that at least a portion of each nano-crystal is in direct contact with the nano-crystal.
  • external medium arranged on a solid support, such as glass slide for example.
  • devices based on the detection of a fluorescent signal are used in particular in the areas based on the detection of chemical or biological reactions (for example biochips).
  • Biochips are a biochemical tool for mass collection of information including nucleic acids (DNA chips) and amino acids (protein chips), antigens and antibodies (immunological sensors). Combined with digital processing techniques harvested information, DNA chips can conduct research (detection, separation, identification, study) to access directly to the DNA.
  • DNA chips can conduct research (detection, separation, identification, study) to access directly to the DNA.
  • the protein chips can detect, identify, separate, study the proteins and determine their activities, functions, interactions, changes over time.
  • the immunological sensors based on binding with an enzyme make it possible to detect antibodies / antigens.
  • a field of application of the invention therefore relates to chips, including biochips, sensors and sensors, which are in the form of a solid support on the surface of which biochemical elements are immobilized. .
  • the present invention finds particular applications in the manufacture of media for sensor or sensor biochips.
  • These include microarray supports compatible with the grafting of nucleic acids (DNA microarrays, RNA biochips), amino acids (protein chips, immunological chips), as well as the cellular biochips used in particular in transfection studies.
  • the invention relates to a support for a biochip, sensor or sensor comprising a substrate whose surface comprises a layer of (nano) material or film according to the invention.
  • the (nano) material comprises at least one inorganic and / or organo-mineral layer, in which at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of said so that at least a part of the nano-crystal is in direct contact with the external medium.
  • the support comprises a substrate on which the (nano) material of the present invention is deposited, in particular in the form of a thin layer or film.
  • It may be any solid material known to those skilled in the art, such as, for example, the support materials used for the manufacture of analytical microsystems and biochips.
  • It can be an organic or inorganic substrate. It may consist for example of a material selected from the group consisting of glass, silica, polycarbonate, nymon, polymethyl methacrylate (PMMA), polystyrene, cycloolefin copolymer (COC), and acrylonitrile polystyrene (SAN).
  • the support is typically a glass slide of comparable size to a microscope slide.
  • the substrate may be previously cleaned in order to improve the adhesion of the (nano) material layer to its surface.
  • This cleaning can be for example a cleaning chemical, possibly followed by a heat treatment.
  • Cleaning techniques in particular those used for the preparation of biochip carriers, sensor and sensor, known to those skilled in the art are usable.
  • any solvent suitable for dusting and / or degreasing the surface of the substrate may be used, preferably without damaging it.
  • Trichlorethylene, acetone, ethyl alcohol, deionized water, acidic or basic solutions, etc. may be mentioned as cleaning solvents, for example.
  • the cleaning may consist of bathing the substrate in one or more of these solvents successively. Cleaning is usually followed by drying.
  • the cleaning may also simply consist of ridding the substrate of its dust with a jet of compressed air.
  • the (nano) material layer may be deposited on the surface of the substrate by any technique known to those skilled in the art for depositing this type of material on a surface.
  • this step can be carried out by:
  • a step (Oa) for preparing an initial mixture for example by mixing in a solvent at least one fluorescent organic or organometallic compound and at least one metal alkoxide, for example of formula (I), in the presence water; eventually ,
  • step (iii) a step of depositing the sol-gel mixture obtained in step (ii) on a support, and (iv) a "spin-coating" centrifugation step to obtain an inorganic and / or organo-mineral layer comprising fluorescent organic organometallic organic nano-crystals on the surface of the support.
  • a sol-gel mixture being prepared by hydrolysis and condensation of salts or alkoxides of metals or metalloids M, preferably in the presence of an organic solvent, this organic solvent can facilitate the deposition and drying of the layer of (nano) material on the substrate.
  • the (nano) material layer according to the invention may be deposited on said surface of the substrate at a thickness of 50 nm to 1000 nm, or even 100 nm to 1000 nm.
  • the minimum thickness is a thickness which advantageously makes it possible to cover the surface of the substrate without leaving holes.
  • the maximum thickness it is generally preferable not to go beyond the micrometer. In fact, if the thickness is greater than one micrometer, problems of cracking / cracking of the layer related to the evaporation of the solvent can occur (internal tensions due to capillary forces, see reference book of Brinker and Scherer [ref I] ).
  • the inorganic and / or organo-mineral layer comprises fluorescent organic or organometallic nanocrystals deposited on the support
  • the latter may be furthermore subjected to a step of dissolving part of a inorganic and / or organo-mineral layer so as to bring out at least a portion of the nanoparticles crystals of the inorganic and / or organo-mineral layer or to increase the surface of the nano-crystals in contact with the external medium.
  • the various embodiments described above for the preparation of the (nano) material according to the invention are applicable to the implementation of supports for chips, sensors or sensors (eg, solvents, fluorescent compounds, metal alkoxides, etc.).
  • one or more types of identical or different probe may be grafted or adsorbed on the part of the nano-crystals emerging from the surface of the inorganic and / or organo-mineral layer.
  • we can apply the various embodiments described above for the (nano) material of the invention i.e., types of probe, modes of connection, etc.).
  • the support according to the invention may comprise a stack of thin dielectric layers forming a Bragg mirror interposed between the substrate and the nanomaterial layer.
  • the reader can refer to the patent documents of Genewave, for example WO 2004/042376 [Ref 5] and WO 2007/045755 [Ref 6]. These are substrates having a Bragg mirror at the grating surface so that the fluorescent signal is completely reflected back to the detector to increase the sensitivity of the system. These substrates are commonly called “amplislides”. This particular configuration may make it possible to increase the exciter field inside the nanomaterial layer.
  • this configuration has the effect of increasing the excitation of luminescent organic or organometallic nano-crystals anchored in the nanomaterial layer, and therefore of increasing the quantity of light emitted into the substrate.
  • a Bragg mirror is a successive stack of several thin dielectric layers of refractive indices n : and n 2 different. The thickness of each of these layers is equal to ⁇ / (4n), with n being able to take the value n : or n 2 .
  • the variable 1 corresponds to the wavelength at which it is desired to have a maximum reflection for the Bragg mirror.
  • the invention relates to the use of the (nano) material according to the invention for the manufacture of media for chips, including supports for biochips, sensors or sensors.
  • the invention relates to a chip, in particular a bio-chip, a sensor or a sensor comprising at least one (nano) material or a film according to the invention.
  • Chips including biochips, sensors and sensors and their various modes of implementation are known to those skilled in the art, have been the subject of numerous reports and publications. For illustrative purposes, reference can be made to (i) D. L. Gerhold, "Better Therapeutics through Microarrays, Nature Genetics, 32, p 547-552 (2002) [Ref 7]; (ii) M.A.
  • oligonucleotides or polynucleotides can be grafted onto or adsorbed onto the luminescent organic or organometallic nanocrystals emerging from the surface of the inorganic and / or organo-mineral layer. (nano) material according to the invention.
  • these oligonucleotides or polynucleotides are known single-stranded oligonucleotides or specific polynucleotides. Their role is to detect complementary labeled targets present in the complex mixture to be analyzed. Indeed, the detection principle used in DNA chips is based on the possibility of pairing DNA strands with their complementary bases.
  • oligonucleotides or polynucleotides are all probes that make it possible to hybridize complementary DNA sequences originating from a biological sample to be analyzed.
  • the organic nano-crystals or organo-metallic luminescent are fluorescent.
  • the chain detection is performed by measuring the corresponding fluorescent signal.
  • the images obtained are digitized and then processed by specific processing algorithms of these data, implemented by computer means.
  • the various embodiments described above for the preparation of the (nano) material and supports for chips, sensors and sensors according to the invention are applicable to the implementation of chips, sensors or sensors (eg, solvents, fluorescent compounds, metal alkoxides, etc.).
  • one or more types of identical or different probes may be grafted or adsorbed on the part of the nano-crystals emerging from the surface of the inorganic and / or organo-mineral layer.
  • the various embodiments described above for the (nano) material of the invention ie, types of probe, modes of connection, etc.).
  • the (nano) material according to the invention can make it possible to obtain a better sensitivity (capacity of detection), and for example to make it possible to win up to 2
  • each nano-crystal may be composed of a very large number of fluorophores (from 10 4 to 10 10 fluorophores per nano-crystal according to its diameter), and
  • the sensitivity can be improved by a factor of 100 or more compared to an isolated fluorophore.
  • This improvement can be observed by measuring the number of moles of fluorophores extinguished in an aqueous suspension of nanocrystals per mole of fluorescence inhibitor.
  • the observed inhibition effect modification of fluorescence decline) is much greater in the case of nano-crystals.
  • the fluorescence chips according to the invention may have a detection capacity (sensitivity) of a target molecule for one hundred fluorescent molecules.
  • This detection capacity, or sensitivity is better measured by means of crystals in solution where the concentrations are better known.
  • An example of this embodiment is given in the Example
  • FIG. 1 represents an example of a "Stern Volmer" plot of Io / I whose slope gives the number of rubrene molecules in the nanoparticle extinguished by each quencher molecule, for example, cibacron blue (CB). In other words, it is the average number of rubrene molecules per fluorescent organic nano-crystal that are extinguished by each quencher molecule.
  • FIG. 2 represents an example of an image by confocal photon microscopy of rubrene nano-crystals in sol-gel matrix derived from TMOS / TMSE precursor.
  • FIG. 3 represents an exemplary image by transmission electron microscopy of rubrene nano-crystals in a sol-gel matrix derived from TMOS / TMSE precursor.
  • FIG. 4 represents an example of images recorded under a microscope at atomic force (AFM) in intermittent mode showing rubrene nanocrystals emerging about 35 nm from the surface of a sol-gel thin layer derived from TMOS / TMSE precursor.
  • FIG. 5 represents an example of images recorded using a near-field atomic force microscope (AFM) showing nanocrystals of rhodamine B emerging from 2 to 3 nm from the surface of a silicate sol-gel thin layer originating from TMOS / GPTMS precursor.
  • FIG. 3 represents an exemplary image by transmission electron microscopy of rubrene nano-crystals in a sol-gel matrix derived from TMOS / TMSE precursor.
  • FIG. 4 represents an example of images recorded under a microscope at atomic force (AFM) in intermittent mode
  • FIG. 6 represents an example of images recorded using a near-field atomic force microscope (AFM) showing auramine 0 nanocrystals emerging from 15 to 20 nm from the surface of a silicate sol-gel thin layer resulting from TMOS / TMSE precursor.
  • AFM near-field atomic force microscope
  • the method of partially dissolving the surface of the sol-gel layers to make the organic nanocrystals emerging according to the invention was applied to different nanocomposite layers in order to illustrate the generic aspect. of the invention. Stripping was carried out on three different sol-gel layers containing different organic nanocrystals. These are detailed in Examples 1 to 3 below.
  • sol-gel mixture is very stable. It may therefore be possible to leave the mixture in storage for several weeks, or even several months if desired. However, several days are sufficient to properly hydrolyze and condensation of starting alkoxides.
  • a volume of 200 .mu.l of the mixture thus obtained is deposited on a slide of microscope glass.
  • the microscope slide is then introduced into a spin coater of the company Suss Microtech type RC8 GYRSET.
  • the sol-gel thin layer is obtained by spin-coating using the following conditions: [0141] rotational speed: 4000 rpm [0142] acceleration: 3000 rpm 2 [0143] duration of the rotation: 10s
  • the thin layer thus obtained is then annealed at 100 ° C. between 24 and 72 hours.
  • the nanocrystals were visualized by confocal optical microscopy (FIG. 2) and transmission electron microscopy (FIG. 3).
  • the nanocomposite thin layer encrusted with rubrene nanocrystals is then subjected to controlled dissolution conditions.
  • the thin layer is etched to expose the surface rubrene nanocrystals by soaking in a dilute NaOH solution (10 -3 M) for 16 hours.
  • the nanocrystals emerging on the surface of the sol-gel layer are shown in FIG. 4.
  • This figure makes it possible to directly view a slide supporting a sol-gel layer of very low roughness (0.5 nm RMS) that supports nanocrystals. emerging which are the signaling function of this new type of detector.
  • the images of FIG. 4 were recorded using an atomic force microscope (AFM) in intermittent mode showing rubrene nanocrystals emerging on the surface of the sol-gel thin layer. These images show that the nanocrystals are in good contact with the external environment but that they remain well anchored in the layer. They are stripped to less than 50% of their diameter, and of the order of 30% maximum.
  • the nanocrystals emerge about 35 nm above the sol-gel layer.
  • the molar proportions in the initial mixture are: 0.4 TMOS + 0.6 GPTMS + 20 MeOH + 8 H 2 O + 0.01 rhodamine B.
  • H 2 O refers to the amount of water introduced with 0.1M HCl solution.
  • the mixture thus obtained was stored for several days to advance the reactions of hydrolysis and condensation of the alkoxides. It is noted that the sol-gel mixture is very stable. It may therefore be possible to leave the mixture in storage for several weeks, or even several months if desired. However, several days are sufficient to properly hydrolyze and condensation of starting alkoxides.
  • a sol-gel thin layer was produced on a microscope slide by spin-coating under spin coating and annealing conditions identical to those of Example 1. [0154] Thus, rhodamine B nanocrystals were obtained. 40 nm in average diameter included in a silicate thin layer 180 nm thick.
  • the blade is then immersed in a solution of NaOH at 1.10 ⁇ 3 mol / L for 4 hours.
  • the nanocrystals emerge after etching of 2 to 3 nm (see FIG. 5).
  • EXAMPLE 3 AURAMINE O NANO-CRYSTALS IN SILICATED THIN-GEL THIN LAYERS FROM PRECURSOR TMOS / TMSE
  • a sol-gel mixture was prepared according to a procedure similar to that of Example 1, using 42.36 mg of auramine microcrystalline powder 0 3.528 mL of THF, 0.257 mL of tetramethylsiloxane (TMOS), 0.222 mL of 1,2-bis (trimethoxysilyl) ethane (TMSE) and 0.219 mL of a 0.1 HCl solution M.
  • the molar proportions in the initial mixture are: 2 TMOS + 1 TMSE + 75 THF + 10 H 2 O + 0.04 auramine 0. (H 2 O refers to the amount of water introduced with 0.1M HCl solution ).
  • the mixture thus obtained was stored for several days (3 or 4 days) to advance the reactions of hydrolysis and condensation of the alkoxides.
  • the sol-gel mixture is stable. It may therefore be possible to leave the mixture in storage for several weeks, or even several months if desired. However, several days are sufficient to properly hydrolyze and condense the starting alkoxides.
  • a sol-gel thin layer was made on a microscope slide by spin-coating under spin coating and annealing conditions identical to those of the previous examples.
  • Auramine 0 nanocrystals of 200 to 300 nm in mean diameter are thus obtained, which are included in a silicate thin film 300 nm thick.
  • An aqueous suspension of nanocrystals is prepared. 10 ml of a solution of rubrene at 1.3 mmolZL in a mixture of ethanol ZTHF 3: 7 is injected with a syringe with a small needle into 100 ml of an aqueous solution of CTACl (cethyltrimethylammonium chloride) at 5 ml. mmolZL. The solution is filtered through a millipore filter of cut size of 50 nm. The concentration of fluorescent molecule is measured by absorption in a quartz cuvette of 1 cm optical path.
  • CTACl cethyltrimethylammonium chloride
  • the fluorescence is measured in a SPEX Fluorolog spectrofluorometer with excitation at 495 nm with 2 nm slots at emission and excitation.
  • the fluorescence intensity is measured for known "quencher” additions (probe molecule which inhibits the fluorescence of the nanocrystals by non-radiative energy transfer between the initially excited nano-crystal and this molecule lying on the surface of the nano-crystal) .
  • a typical quencher is cibachron blue. Another is methylene blue. Both are dissolved at 1 mmol / L in water. It is shown in FIG.

Abstract

The present invention relates to a (nano)material comprising at least one inorganic and/or organomineral layer, integrated into which is at least one type of organic or organometallic fluorescent nanocrystal that emerges from the surface of said layer so that at least one part of the fluorescent nanocrystals is in direct contact with the outside environment. These emerging nanocrystals constitute, due to their fluorescence, a signalling function, and they may thus be functionalized by grafting a biomolecule bearing a probe function, such as a half-strand of DNA, to thus produce a biochip, biocaptor or biosensor comprising such a (nano)material, a support for a biochip, biocaptor or biosensor, and a process for preparing such a nanomaterial.

Description

NANOCRISTAUX ORGANIQUES LUMINESCENTS POUR LA RÉALISATION DELUMINESCENT ORGANIC NANOCRYSTALS FOR THE PRODUCTION OF
CAPTEURS BIOLOGIQUESBIOLOGICAL SENSORS
Priorité [0001] La présente invention revendique le droit de priorité à la demande de brevet français N 07/53947 déposée le 21 mars 2007. Le contenu de cette demande est incorporé, par la présente, par référence, dans son intégralité.Priority [0001] The present invention claims the right of priority to the French patent application N 07/53947 filed March 21, 2007. The content of this application is hereby incorporated by reference in its entirety.
Domaine techniqueTechnical area
[0002] La présente invention concerne le domaine des matériaux à propriétés optiques, et en particulier des capteurs et des senseurs, notamment biologiques. Tout particulièrement il peut s'agir de puces, notamment de biopuces, en particulier à fluorescence.The present invention relates to the field of materials with optical properties, and in particular sensors and sensors, including biological. In particular, it may be chips, especially biochips, in particular fluorescence.
[0003] Dans la description ci-dessous, les références entre crochets ( [ ] ) renvoient à la liste des références présentées après les exemples.In the description below, the references in square brackets ([]) refer to the list of references presented after the examples.
Etat de la techniqueState of the art
[0004] Les biopuces sont des outils majeurs dans la recherche clinique, en particulier pour le développement de tests diagnostics et pronostics, ainsi que dans la recherche de nouvelles procédures thérapeutiques.[0004] Biochips are major tools in clinical research, in particular for the development of diagnostic tests and prognoses, as well as in the search for new therapeutic procedures.
[0005] Les biopuces les plus couramment utilisées sont les biopuces à fluorescence. Cependant, celles-ci peuvent présenter une sensibilité insuffisante pour procéder à l'analyse d'échantillons précieux et chers, fragiles et/ou compliquées à fabriquer. Les biopuces peuvent encore être insuffisamment transparentes, en particulier dans certaines longueurs d'ondes, comme l ' Infra-Rouge. Elles peuvent également être insuffisamment stables, et par exemple ne pas présenter une résistance chimique ou temporelle satisfaisante.The most commonly used biochips are fluorescence biochips. However, these may have insufficient sensitivity to analyze precious and expensive samples, fragile and / or complicated to manufacture. Biochips may still be insufficiently transparent, especially in certain wavelengths, such as Infra-Red. They may also be insufficiently stable, for example not have satisfactory chemical or temporal resistance.
[0006] Tout particulièrement les procédés permettant de préparer les matériaux permettant de fabriquer des puces, et notamment des biopuces, ou de préparer les puces sont généralement onéreux, difficiles à être mis en place de manière « industrielle » (en particulier pour préparer un grand nombre de puces en même temps et/ou en une seule opération), délicats et/ou complexes à mettre en œuvre.[0006] In particular, the processes making it possible to prepare the materials making it possible to manufacture chips, and in particular biochips, or to prepare the chips, are generally expensive, difficult to implement in an "industrial" manner (in particular to prepare a large number of chips). number of chips at the same time and / or in a single operation), delicate and / or complex to implement.
[0007] II existe donc un besoin pour des puces qui permettent de résoudre en tout ou partie les problèmes évoqués ci-dessus, notamment en termes de résistance, en particulier thermique, mécanique, chimique et/ou photochimique ; et/ou des biopuces qui permettent de présenter de bonnes propriétés optiques, comme la fluorescence, une bonne stabilité, une grande gamme de transparence, ainsi qu'un procédé de fabrication et de mise en forme aisés, une faible diffusion et une bonne stabilité.There is therefore a need for chips that can solve all or part of the problems mentioned above, particularly in terms of resistance, in particular thermal, mechanical, chemical and / or photochemical; and / or biochips which make it possible to present good optical properties, such as fluorescence, good stability, a wide range of transparency, as well as an easy manufacturing and shaping process, low diffusion and good stability.
[0008] II existe d'autre part également un besoin pour un (nano)matériau permettant de préparer des puces permettant de résoudre en tout ou partie les problèmes évoqués ci-dessus. [0009] II existe encore un besoin pour un procédé de préparation de ce (nano)matériau permettant de résoudre en tout ou partie les problèmes évoqués ci-dessus.On the other hand, there is also a need for a (nano) material for preparing chips to solve all or part of the problems mentioned above. There is still a need for a process for preparing this (nano) material to solve all or part of the problems mentioned above.
Description de l'invention [0010] Ainsi, selon un premier aspect, l'invention a pour objet un (nano)matériau comprenant au moins une couche inorganique et/ou organo-minérale, dans laquelle est intégré au moins un type de nano-cristal luminescent dont au moins une partie est en contact direct avec le milieu extérieur.DESCRIPTION OF THE INVENTION [0010] Thus, according to a first aspect, the subject of the invention is a (nano) material comprising at least one inorganic and / or organo-mineral layer, in which at least one type of nanoparticles is integrated. luminescent crystal at least a portion is in direct contact with the external environment.
[0011] Avantageusement, les nano-cristaux organiques ou organo-métalliques luminescents sont des nano-cristaux fluorescents.Advantageously, the organic nano-crystals or organo-metallic luminescent are fluorescent nano-crystals.
[0012] Dans un mode de réalisation, le (nano)matériau comprend au moins une couche inorganique et/ou organo- minérale, dans laquelle est intégré au moins un type de nano-cristal fluorescent organique ou organo-métallique émergeant de la surface de ladite couche de sorte qu'au moins une partie du nano-cristal est en contact direct avec le milieu extérieur.In one embodiment, the (nano) material comprises at least one inorganic and / or organo-mineral layer, in which is integrated at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of the said layer so that at least a portion of the nano-crystal is in direct contact with the external medium.
[0013] Par « (nano)matériau », on entend au sens de la présente invention un matériau ou un nanomatériau tel que défini ci-dessous.By "(nano) material" is meant in the sense of the present invention a material or a nanomaterial as defined below.
[0014] Par « nanomatériau », on entend au sens de la présente invention un matériau de dimension nanométrique, c'est-à-dire un matériau dont au moins une des dimensions est à l'échelle nanométrique. Autrement dit, il s'agit d'un matériau dont la taille nanométrique dans au moins une des dimensions de l'espace. Par exemple, la taille du matériau dans au moins une des dimensions de l'espace est comprise entre 1 et 500nm, de préférence entre 1 et 100 nm. Par exemple, il peut s'agir d'une couche mince. [0015] Par « couche organo-minérale », on entend au sens de la présente invention une matrice comprenant au moins un composé minéral et au moins un composé organique. Ainsi, la couche organo-minérale peut être une couche sol-gel, par exemple de variété Silicate, Titanate, Zirconate, Stannate, Borate, Aluminate ou Yttriate, et comprenant au moins des nanocristaux organiques ou organo-métalliques. [0016] La couche sol-gel peut en particulier être constituée majoritairement de silicate/oxyde de silicium amorphe, qui peut résulter de l'hydrolyse et polycondensation des alkoxydes de silicium inclus dans la solution initiale.For the purposes of the present invention, the term "nanomaterial" means a material of nanometric size, that is to say a material of which at least one of the dimensions is at the nanoscale. In other words, it is a material whose nanometric size in at least one of the dimensions of the space. For example, the size of the material in at least one of the dimensions of the space is between 1 and 500 nm, preferably between 1 and 100 nm. For example, it may be a thin layer. For the purposes of the present invention, the term "organo-mineral layer" means a matrix comprising at least one mineral compound and at least one organic compound. Thus, the organomineral layer may be a sol-gel layer, for example of the Silicate, Titanate, Zirconate, Stannate, Borate, Aluminate or Yttriate variety, and comprising at least organic or organometallic nanocrystals. The sol-gel layer may in particular consist mainly of silicate / amorphous silicon oxide, which may result from the hydrolysis and polycondensation of the silicon alkoxides included in the initial solution.
[0017] Cependant la couche sol-gel, ou le réseau polymérique, peut contenir également des radicaux carbonés alkyles, aryles et/ou aralkyles.However, the sol-gel layer, or the polymeric network, may also contain alkyl, aryl and / or aralkyl carbon radicals.
[0018] La couche inorganique et/ou organo-minérale peut tout particulièrement comprendre au moins une couche sol- gel. [0019] Cette couche sol-gel peut particulièrement être une de variété silicate ou métalliques, en particulier accessible à partir de précurseurs alcoxydes de silicium, d'alcoxydes de titane ou d'autres métaux, en particulier choisis parmi Titane, Zirconium, Étain, Bore, Aluminium et Yttrium.The inorganic and / or organo-mineral layer may especially comprise at least one sol-gel layer. This sol-gel layer may especially be one of silicate or metal variety, in particular accessible from precursor alkoxides of silicon, titanium alkoxides or other metals, in particular selected from titanium, zirconium, tin, Boron, Aluminum and Yttrium.
[0020] Selon la finalité de la matrice (balance hydrophile-hydrophobe, amélioration de la cristallinité, taille des nanocristaux, ...) les différents types d'alcoxydes pourront être utilisés à 100 % ou sous forme de mélange en proportions variables.Depending on the purpose of the matrix (hydrophilic-hydrophobic balance, improvement of crystallinity, size of nanocrystals, ...) the different types of alkoxides can be used at 100% or as a mixture in varying proportions.
[0021] La chimie sol-gel et ses divers modes de mise en oeuvre sont connus de l'homme de métier, ont fait l'objet de nombreux rapports et publications, et ne seront pas développés dans le cadre du présent brevet. On citera par exemple, l'ouvrage de CJ. Brinker et G.W. Scherer, Sol-Gel Science, The Physics and Chemistry of SoIGeI Processing, Académie Press, New York, 1990 [ref I]. On peut également citer d'autres articles de synthèse tels que D. Avnir, V.R. Kaufman and R. Reisfeld, J. Non. Cryst. Solids, 1985, 74, 395-406 [ref 2]; et C. Sanchez and F. Ribot, New J. Chem. , 1994, 18, 1007-1047 [ref 3].Sol-gel chemistry and its various modes of implementation are known to those skilled in the art, have been the subject of numerous reports and publications, and will not be developed in the context of this patent. For example, CJ's book. Brinker and G.W. Scherer, Sol-Gel Science, The Physics and Chemistry of SoIngI Processing, Academy Press, New York, 1990 [ref I]. Other review articles such as D. Avnir, V.R. Kaufman and R. Reisfeld, J. No. Cryst. Solids, 1985, 74, 395-406 [ref 2]; and C. Sanchez and F. Ribot, New J. Chem. , 1994, 18, 1007-1047 [ref 3].
[0022] De manière générale, la couche inorganique et/ou organo-minérale comprenant une couche sol-gel peut être obtenue à partir d'un mélange sol-gel. Avantageusement, le mélange sol-gel contient au moins un composé organique ou organo-métallique fluorescent .Avantageusement , ce composé organique ou organo-métallique fluorescent initialement dissous dans le mélange sol-gel est cristallisé au sein de la matrice sol-gel pour former des nano-cristaux organiques ou organo-métalliques fluorescents. La cristallisation peut être effectuée par séchage du mélange sol-gel contenant le composé organique ou organo-métallique fluorescent. La vitesse du séchage peut être ajustée pour favoriser la formation de nano-cristaux.In general, the inorganic and / or organo-mineral layer comprising a sol-gel layer may be obtained from a sol-gel mixture. Advantageously, the sol-gel mixture contains at least one fluorescent organic or organometallic compound. Advantageously, this fluorescent organic or organometallic compound initially dissolved in the sol-gel mixture is crystallized in the sol-gel matrix to form nano-crystals. organic or organo-metallic fluorescent. The crystallization can be carried out by drying the sol-gel mixture containing the organic or organo-metallic fluorescent compound. The speed of drying can be adjusted to promote the formation of nano-crystals.
[0023] On entend par mélange « sol-gel » au sens de la présente invention un mélange comprenant initialement au moins un alcoxyde métallique en présence d'eau, au moins un solvant organique et au moins un composé organique ou organo-métallique fluorescent, ledit mélange étant initialement sous la forme d'une solution et produisant un gel par réaction d'hydrolyse et de polycondensation. En effet, la présence d'eau permet d'initier des réactions d'hydrolyse et de condensation des alcoxydes métalliques formant un réseau inorganique conduisant à des gels puis des xérogels. Ces xérogels conduisent à un réseau solide inorganique structuré d'oxydes métalliques. La chimie sol- gel est par exemple citée dans le brevet français FR 2 853 307 [ref 4]. [0024] La couche sol-gel est susceptible d'être obtenue à partir de tout alcoxyde métallique connu de l'homme du métier compatible avec la chimie sol-gel.For the purposes of the present invention, the term "sol-gel" is understood to mean a mixture initially comprising at least one metal alkoxide in the presence of water, at least one organic solvent and at least one fluorescent organic or organo-metallic compound, said mixture being initially in the form of a solution and producing a gel by hydrolysis and polycondensation reaction. In fact, the presence of water makes it possible to initiate hydrolysis and condensation reactions of metal alkoxides forming an inorganic network, leading to gels and then xerogels. These xerogels lead to a structured inorganic solid network of metal oxides. Sol-gel chemistry is for example cited in French patent FR 2,853,307 [ref 4]. The sol-gel layer can be obtained from any metal alkoxide known to those skilled in the art compatible with sol-gel chemistry.
[0025] Ainsi, dans un mode de réalisation, la couche sol-gel est susceptible d'être obtenue à partir d'un alcoxyde métallique de formule (I) :Thus, in one embodiment, the sol-gel layer can be obtained from a metal alkoxide of formula (I):
R1 xM(0R2)y R 1 x M (OR 2 ) y
(I) dans laquelle : M est un métal choisi dans le groupe comprenant Si, Ti, Zr, Sn, B, Al et Y, par exemple Si. x est un nombre entier allant de 0 à 2 , par exemple 0 ou 1 ; y est un nombre entier allant de 1 à 6 , par exemple 3 ou 4 ; x+y correspond au nombre de coordination du métal M ;(I) in which: M is a metal selected from the group consisting of Si, Ti, Zr, Sn, B, Al and Y, for example Si x is an integer from 0 to 2, for example 0 or 1; y is an integer ranging from 1 to 6, for example 3 or 4; x + y corresponds to the coordination number of the metal M;
R1 et R2 représentent indépendamment un radical organique compatible avec la chimie sol-gel.R 1 and R 2 independently represent an organic radical compatible with sol-gel chemistry.
[0026] L'homme du métier saura sélectionner R1 et R2 à la lumière des ouvrages précités et de ses connaissances générales dans le domaine de la chimie sol-gel. Les exemples donnés ci-dessous sont donnés purement à titre illustratif et ne sont en aucun cas limitatifs. [0027] Par exemple, R1 et R2 peuvent représenter indépendamment un radical carboné alkyle, aryle ou aralkyle. [0028] Les radicaux alkyles peuvent être linéaires ou ramifiés, cycliques ou acycliques, saturés ou insaturés, et éventuellement comprendre ou porter des hétéroatomes, notamment 0, N, S, B et P. Il peut par exemple s'agir de C^^alkyle; C2_15alcenyle; C2_15alcynyle; C3_15cycloalkyle. [0029] Les radicaux alkyles peuvent comprendre de 1 à 15 atomes de carbones, et notamment être choisis parmi les radicaux méthyle, éthyle, propyle, butyle, pentyle, hexyle, heptyle, octyle, nonyle, décyle, undécyle et dodécyle. Selon l'invention, le radical alkyle peut comprendre 1 à 15 atomes de carbone, par exemple 1 à 10 atomes de carbone, par exemple 1 à 6 atomes de carbone.The skilled person will select R 1 and R 2 in the light of the above works and his general knowledge in the field of sol-gel chemistry. The examples given below are purely illustrative and are in no way limiting. For example, R 1 and R 2 may independently represent an alkyl, aryl or aralkyl carbon radical. The alkyl radicals may be linear or branched, cyclic or acyclic, saturated or unsaturated, and optionally include or carry heteroatoms, especially O, N, S, B and P. It may for example be C ^^ alkyl; C 2 _ 15 alkenyl; C 2 _ 15 alkynyl; C 3 -C 15 cycloalkyl. The alkyl radicals can comprise from 1 to 15 carbon atoms, and in particular be chosen from methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl and dodecyl radicals. According to the invention, the alkyl radical can comprise 1 to 15 carbon atoms, for example 1 to 10 carbon atoms, for example 1 to 6 carbon atoms.
[0030] On notera que les radicaux alkyles peuvent comprendre ou porter un ou plusieurs hétéroatomes, notamment choisis dans le groupe comprenant l'oxygène, le soufre, l'azote, le phosphore et le bore. Autrement dit, les composés de formule (I) où R1 et/ou R2 est un radical alkyle englobent les composés de formule (I) où R1 et/ou R2 est un radical hétéroalkyle. Il peut par exemple s'agir d'un hétéroalkyle, d'un hétéroalcenyle, d'un hétéroalcynyle, d'un hétérocycle, d'un alkoxy, d'un alkylthio, d'une alkylamine, d'un alkylamide, d'une alkylimine, d'un alkylimide, d'un alkylester, d'un alkyléther, etc. Selon l'invention, le radical hétéroalkyle peut comprendre 1 à 15 atomes de carbone, par exemple 1 à 10 atomes de carbone, par exemple 1 à 6 atomes de carbone. [0031] On entend par « aryle » au sens de la présente invention, un radical comprenant au moins un cycle satisfaisant la règle d'aromaticité de Hϋckel. Ledit aryle peut être mono- ou polycyclique, fusionné ou non. Les radicaux aryles peuvent comprendre de 4 à 15 atomes de carbone, en particulier de 6 à 10 atomes de carbone, et éventuellement comprendre des hétéroatomes, notamment 0, N, S. Parmi les radicaux aryles, on peut citer les phényles, naphtyles et pyridinyles.It will be noted that the alkyl radicals may comprise or carry one or more heteroatoms, in particular chosen from the group comprising oxygen, sulfur, nitrogen, phosphorus and boron. In other words, the compounds of formula (I) in which R 1 and / or R 2 is an alkyl radical include the compounds of formula (I) in which R 1 and / or R 2 is a heteroalkyl radical. It may for example be a heteroalkyl, a heteroalkenyl, a heteroalkynyl, a heterocycle, an alkoxy, an alkylthio, an alkylamine, an alkylamide, a alkylimine, alkylimide, alkyl ester, alkyl ether, etc. According to the invention, the heteroalkyl radical may comprise 1 to 15 carbon atoms, for example 1 to 10 carbon atoms, for example 1 to 6 carbon atoms. For the purposes of the present invention, the term "aryl" means a radical comprising at least one ring satisfying Hekel's aromaticity rule. Said aryl may be mono- or polycyclic, fused or not. The aryl radicals may comprise from 4 to 15 carbon atoms, in particular from 6 to 10 carbon atoms, and optionally include heteroatoms, in particular O, N, and S. Among the aryl radicals, mention may be made of phenyls, naphthyls and pyridyls. .
[0032] On notera que les radicaux aryles peuvent comprendre ou porter un ou plusieurs hétéroatomes, notamment choisis dans le groupe comprenant l'oxygène, le soufre, l'azote et le phosphore. Autrement dit, les composés de formule (I) où R1 et/ou R2 et un radical aryle englobent les composés de formule (I) où R1 et/ou R2 est un radical hétéroaryle, c'est-à-dire un groupe cyclique ou polycyclique hydrocarboné aromatique dans lequel au moins un atome de carbone est remplacé par un hétéroatome, notamment choisi dans le groupe comprenant l'oxygène, le soufre et l'azote.It will be noted that the aryl radicals may comprise or carry one or more heteroatoms, in particular chosen from the group comprising oxygen, sulfur, nitrogen and phosphorus. In other words, the compounds of formula (I) wherein R 1 and / or R 2 and an aryl radical include the compounds of formula (I) wherein R 1 and / or R 2 is a heteroaryl radical, that is to say an aromatic hydrocarbon ring or polycyclic group in which at least one carbon atom is replaced by a heteroatom, in particular chosen from the group comprising oxygen, sulfur and nitrogen.
[0033] On entend par « aralkyle » au sens de la présente invention, un radical alkyle substitué par un radical aryle, les termes « alkyle » et « aryle » étant tels que définis ci-dessus. Le point d'attachement du radical « aralkyle » au reste de la molécule (c'est-à-dire au métal M dans la formule (I)) est au niveau du groupe alkyle. Les radicaux aralkyles peuvent comprendre de 4 à 30 atomes de carbone, en particulier de 6 à 20 atomes de carbone, et éventuellement comprendre ou être substitués par des radicaux alkyles et/ou des hétéroatomes, notamment 0, N, S et P. Parmi les radicaux aralkyles, on peut citer benzyle et tolyle.For the purposes of the present invention, the term "aralkyl" means an alkyl radical substituted with a radical. aryl, the terms "alkyl" and "aryl" being as defined above. The point of attachment of the "aralkyl" radical to the remainder of the molecule (i.e., to the metal M in formula (I)) is at the alkyl group. The aralkyl radicals can comprise from 4 to 30 carbon atoms, in particular from 6 to 20 carbon atoms, and optionally comprise or be substituted by alkyl radicals and / or heteroatoms, in particular O, N, S and P. Among the Aralkyl radicals include benzyl and tolyl.
[0034] Dans un mode de réalisation particulier, R1 et R2 peuvent représenter indépendamment un radical alkyle en C1 à C15, hétéroalkyle C1 à C15, aryle en C6 à C25, hétéroaryle en C4 à C25, ou aralkyle en C6 à C20; les radicaux R1 et R2 étant indépendamment éventuellement substitués par un ou plusieurs groupes R indépendamment choisis dans le groupe comprenant un radical alkyle en C1 à C10, hétéroalkyle C1 à C10; aryle en C6 à C10 ou hétéroaryle en C4 à C10 ; F ; Cl ; Br; I; -NO2 ; -CN ; ou une fonction —GRG1 dans laquelle G est -0-, -S-, -NRG2-, -C(=0)-, -C (=0)0-, -C(=0)NRG2-, où chaque occurrence de RG1, RG2 et RG3 est indépendamment des autres occurrences de RG1 un atome d'hydrogène, ou un radical alkyle en C1 à C10, hétéroalkyle C1 à C10; aryle en C6 à C10 ou hétéroaryle en C4 à C10 ; ou bien, lorsque G représente -NRG2-, RG1 et RG2 conjointement avec l'atome d'azote auquel ils sont liés forment un hétérocycle ou un hétéroaryle éventuellement substitué.[0034] In a particular embodiment, R 1 and R 2 may independently represent an alkyl radical C 1 -C 15 heteroalkyl, C 1 -C 15 aryl, C 6 -C 25 heteroaryl, C 4 -C 25 or C 6 -C 20 aralkyl; R 1 and R 2 being independently optionally substituted by one or more R groups independently selected from the group comprising an alkyl radical C 1 -C 10 heteroalkyl, C 1 -C 10; C 6 -C 10 aryl or C 4 -C 10 heteroaryl; F; Cl; Br; I; -NO 2 ; -CN; or a function -GR G1 in which G is -O-, -S-, -NR G2 -, -C (= O) -, -C (= O) O-, -C (= O) NR G2 -, wherein each occurrence of R G1, R G2 and R G3 is, independently of other occurrences of R G1 is hydrogen or an alkyl radical C 1 -C 10 heteroalkyl, C 1 -C 10; C 6 -C 10 aryl or C 4 -C 10 heteroaryl; or when G is -NR G2 -, R G1 and R G2 together with the nitrogen atom to which they are attached form an optionally substituted heterocycle or heteroaryl.
[0035] Les radicaux alkyles, aryles et/ou aralkyles peuvent être liés directement aux atomes de silicium, titanium, zirconium, étain, bore, aluminium, yttrium, notamment par une liaison covalente. On peut alors parler de matériaux hybrides organo-mineraux, ainsi que par exemple les matériaux connus dans le domaine des lentilles de contact obtenues par sol-gel.The alkyl, aryl and / or aralkyl radicals may be bonded directly to the atoms of silicon, titanium, zirconium, tin, boron, aluminum, yttrium, in particular by a covalent bond. We can then speak of hybrid organic-mineral materials, as well as by examples are known materials in the field of contact lenses obtained by sol-gel.
[0036] La couche sol-gel peut comprendre uniquement du silicium, ou du silicium et du Titane, du Zirconium et/ou de l'Étain. Cette couche sol-gel peut notamment être obtenue en utilisant d'autres types d'alkoxydes métalliques, comme des alkoxydes de Titane, Zirconium et/ouThe sol-gel layer may comprise only silicon, or silicon and titanium, zirconium and / or tin. This sol-gel layer can in particular be obtained by using other types of metal alkoxides, such as titanium alkoxides, zirconium and / or
Étain.Tin.
[0037] Dès le mélange sol-gel constitué, les alcoxydes métalliques réagissent via des réactions d'hydrolyse et de condensation, conduisant à des « polymères inorganiques ».Once the sol-gel mixture constituted, the metal alkoxides react via hydrolysis and condensation reactions, leading to "inorganic polymers".
Dans le domaine de la chimie sol-gel, le terme « polymère inorganique » est utilisé par analogie avec les polymères organiques. A titre illustratif, les réactions suivantes d'hydrolyse (1) et de condensation (2) des alcoxydes métalliques peuvent intervenir.In the field of sol-gel chemistry, the term "inorganic polymer" is used in analogy with organic polymers. By way of illustration, the following reactions of hydrolysis (1) and condensation (2) of metal alkoxides can take place.
( 1 ) R1 xM(OR2)y + H2O ^ R1 x(OR2)y-1 M(OH) + HOR2 (1) R 1 x M (OR 2 ) y + H 2 O 2 R 1 x (OR 2 ) y-1 M (OH) + HOR 2
( 2 ) 2 R1 x(OR2)y-1M(OH) ^ R1 x(OR2)y-1 M-O-M(OR2)y-1 R1 X + H2O où R1, R2, x et y sont tels que définis précédemment. [0038] Avantageusement, le mélange sol-gel du procédé de l'invention peut être homogène et stable chimiquement et mécaniquement. Par exemple il peut être stable pendant plusieurs mois voire plusieurs années ce qui présente un fort avantage pour la mise en œuvre pratique de la présente invention. (2) 2 R 1 x (OR 2 ) y-1 M (OH) 2 R 1 x (OR 2 ) y-1 MOM (OR 2 ) y-1 R 1 X + H 2 O where R 1 , R 2 , x and y are as defined above. Advantageously, the sol-gel mixture of the process of the invention can be homogeneous and stable chemically and mechanically. For example, it can be stable for several months or even years, which presents a great advantage for the practical implementation of the present invention.
[0039] On entend par « alcoxydes métalliques », également appelés « précurseurs sol-gel » au sens de la présente invention, tout composé métallique de formule (I). Selon l'invention, on peut utiliser tout alcoxyde métallique connu de l'homme du métier dans la mesure où l'on peut obtenir un réseau sol-gel polymérique interconnecté. L'homme du métier pourra notamment utiliser tout alcoxyde métallique propre à la chimie sol-gel connu. Pour cela, le lecteur peut notamment se référer au livre de référence CJ. Brinker et G.W. Scherer, Sol-Gel Science, The Physics and Chemistry of SoIGeI Processing, Académie Press, New York, 1990 [ref I]. Voir également ref 2 et 3. [0040] On entend par « solvant » au sens de la présente invention tout solvant connu de l'homme du métier compatible avec le procédé de mise en oeuvre de la présente invention, c'est-à-dire pouvant rendre miscibles les alcoxydes métalliques et l'eau d'hydrolyse afin d'obtenir des solutions homogènes, notamment propres à leur mise en forme en couche mince ou sous forme de film. Avantageusement, le solvant permet dissoudre totalement le composé fluorescent. [0041] Le solvant peut être par exemple choisi dans le groupe comprenant les alcools RS-OH où Rs est un radical alkyle en C1 à C6, les cétones RS1-C(=O) -Rs2, les éthers Rsl- 0-Rs2, où Rsl et Rs2 sont indépendamment des radicaux alkyles en C1 à C6, le tétrahydrofurane, l 'acétonitrile, le diméthylformamide, le toluène, le diméthylsulfoxyde, le dioxane, l'acétone, l'acide acétique, l'acide formique, le dichlorométhane, le chloroforme, le dichloroéthane, l'acétate d'éthyle, le diéthyléther ou un mélange de ceux- ci. [0042] Selon l'invention, l'eau présente dans le mélange sol-gel peut être introduite initialement en faible quantité, celle-ci étant suffisante pour initier les réactions conduisant à la polycondensation du réseau inorganique, ou en quantité supérieure à la quantité initiale d'alcoxyde métallique. Par exemple le mélange sol- gel peut comprendre initialement un pourcentage molaire d'eau par rapport au nombre de fonctions alcoxydes (—OR2) de 10 à 400%, de préférence de 20 à 100%, de manière plus préférée de 50 à 100 %. [0043] Les alcoxydes de silicium précurseurs de silicates peuvent être choisis, par exemple, dans le groupe comprenant le tétraméthoxysilane (TMOS, Si(OCH3)4), le tétraéthoxysilane (TEOS, Si(OC2H5)4), le méthyltriméthoxysilane (MTMOS, CH3Si(OCH3)3) , le méthyltriéthoxysilane (MTEOS, CH3Si(OC2H5) 3) , l'éthyltriéthoxysilane (ETEOS, C2H5Si(OC2H5) 3) , le 1,2- bis(triméthoxysilyl)ethane (TMSE), le 3- glycidoxypropyl ) trimethoxysilane (GPTMS), ou un mélange de ceux-ci.The term "metal alkoxides", also known as "sol-gel precursors" within the meaning of the present invention, any metal compound of formula (I). According to the invention, any metal alkoxide known to those skilled in the art can be used insofar as an interconnected polymeric sol-gel network can be obtained. Those skilled in the art may in particular use any metal alkoxide specific to known sol-gel chemistry. For this purpose, the reader may in particular refer to the reference book CJ. Brinker and GW Scherer, Sol-Gel Science, The Physics and Chemistry of SoIngI Processing, Academy Press, New York, 1990 [ref I]. See also ref 2 and 3. For the purposes of the present invention, the term "solvent" means any solvent known to those skilled in the art compatible with the process for implementing the present invention, that is to say to say that can be miscible metal alkoxides and hydrolysis water to obtain homogeneous solutions, especially suitable for their shaping thin film or film. Advantageously, the solvent makes it possible to completely dissolve the fluorescent compound. The solvent may for example be chosen from the group comprising the alcohols R S -OH where R s is a C 1 to C 6 alkyl radical, the ketones R S 1 -C (= O) -R s 2 , the ethers R sl -O -R s2 , where R sl and R s2 are independently C 1 -C 6 alkyl radicals, tetrahydrofuran, acetonitrile, dimethylformamide, toluene, dimethylsulfoxide, dioxane, acetone, acetic acid, formic acid, dichloromethane, chloroform, dichloroethane, ethyl acetate, diethyl ether or a mixture thereof. According to the invention, the water present in the sol-gel mixture can be introduced initially in a small quantity, this being sufficient to initiate the reactions leading to the polycondensation of the inorganic network, or in a quantity greater than the quantity initial metal alkoxide. For example, the sol-gel mixture may initially comprise a molar percentage of water relative to the number of alkoxide functional groups (-OR 2 ) of from 10 to 400%, preferably from 20 to 100%, more preferably from 50 to 100 %. Silicate precursor alkoxides may be chosen, for example, from the group comprising tetramethoxysilane (TMOS, Si (OCH 3 ) 4 ), tetraethoxysilane (TEOS, Si (OC 2 H 5 ) 4 ), methyltrimethoxysilane (MTMOS, CH 3 Si (OCH 3 ) 3 ), methyltriethoxysilane (MTEOS, CH 3 Si (OC 2 H 5 ) 3 ), ethyltriethoxysilane (ETEOS, C 2 H 5 Si (OC 2 H 5 ) 3 ) 1,2-bis (trimethoxysilyl) ethane (TMSE), 3-glycidoxypropyl) trimethoxysilane (GPTMS), or a mixture thereof.
[0044] Pour les alcoxydes métalliques précurseurs de titanates, zirconates, stannates, borate, aluminates et yttriates, le lecteur pourra se référer par exemple à CJ. Brinker et G.W. Scherer, Sol-Gel Science, The Physics and Chemistry of SoIGeI Processing, Académie Press, New York, 1990 [ref I]. Voir également ref 2 et 3.For metal alkoxides precursors of titanates, zirconates, stannates, borate, aluminates and yttriates, the reader can refer for example to CJ. Brinker and G.W. Scherer, Sol-Gel Science, The Physics and Chemistry of SoIngI Processing, Academy Press, New York, 1990 [ref I]. See also ref 2 and 3.
[0045] Dans le cas d'une matrice obtenue par mélange équimoléculaire MTMOS et TMOS, les substituants méthyles non-liants de MTMOS peuvent couvrir, au moins en partie, les pores du gel et favoriser l'agrégation des nano- cristaux, tandis que les groupes silanols peuvent former des liaisons, notamment hydrogènes, avec les molécules organiques .In the case of a matrix obtained by equimolecular mixing MTMOS and TMOS, the non-linking methyl substituents of MTMOS can cover, at least in part, the pores of the gel and promote the aggregation of the nano-crystals, while the silanol groups can form bonds, in particular hydrogen bonds, with the organic molecules.
[0046] Tout particulièrement la couche inorganique et/ou organo-minérale, en particulier la couche sol-gel, se présente sous la forme d'un film.In particular, the inorganic and / or organo-mineral layer, in particular the sol-gel layer, is in the form of a film.
[0047] Par « film », on peut entendre au sens de la présente invention une structure dans laquelle l'épaisseur est de l'ordre de quelques centaines de nanomètres ou de l'ordre du micromètre, et en particulier une épaisseur allant de 50 nm à 1500 nm, en particulier de 80 à 1200 nm, voire de 100 à 1000 nm. [0048] Tout particulièrement, ce film présente une largeur et/ou une longueur de 1 mm à 10 cm, en particulier de 5 mm à 5 cm.By "film" can be understood in the sense of the present invention a structure in which the thickness is of the order of a few hundred nanometers or the order of a micrometer, and in particular a thickness of 50 nm at 1500 nm, in particular from 80 to 1200 nm, or even from 100 to 1000 nm. In particular, this film has a width and / or length of 1 mm to 10 cm, in particular 5 mm to 5 cm.
[0049] Les nano-cristaux peuvent être fluorescents, tout particulièrement ils absorbent et ré-émettent de la lumière dans les longueurs d'onde du visible ou du proche infrarouge.The nano-crystals may be fluorescent, especially they absorb and re-emit light in the wavelengths of the visible or near infrared.
[0050] La luminescence peut se répandre sur un ensemble de molécules luminescentes, ou chromophores, par exemple allant de 103 à 1010, notamment de 103 à 109, en particulier de 104 à 108, voire de 105 à 107 molécules par nano- cristal.The luminescence may spread over a set of luminescent molecules, or chromophores, for example ranging from 10 3 to 10 10 , in particular from 10 3 to 10 9 , in particular from 10 4 to 10 8 , or even from 10 5 to 10 9. 10 7 molecules per nano-crystal.
[0051] Tout particulièrement, les nano-cristaux émettent une fluorescence, qui est dispersée sur l'ensemble des fluorophores et qui est éteinte ou désactivée (« quenched ») simultanément. Par « simultanément », on entend de 0,001 à 10 ns, en particulier de 0,01 à 5 ns, en particulier de 0,1 à 2 ns, voire de 0,5 à 1,5 ns . [0052] On entend par composé ou nanocristal « fluorescent » au sens de la présente invention, un composé ou un nanocristal ayant la propriété d'émettre, suite à une excitation lumineuse, un rayonnement électromagnétique dans le domaine de la lumière visible ou dans le proche infra-rouge (IR)- Le signal lumineux d'excitation peut provenir d'une source lumineuse telle qu'un laser, une lampe à arc ou des diodes électroluminescentes (LED) . Par exemple, le composé ou le nanocristal fluorescent peut émettre un rayonnement électromagnétique aux longueurs d'onde de 400 à 1200 nm, qui correspond à la fenêtre de relative transparence des tissus vivants. Par exemple, le composé ou le nanocristal fluorescent peut émettre un rayonnement électromagnétique aux longueurs d'onde de 400 à 1000 nm, par exemple de 550 à 800 nm, qui correspond à une fluorescence dans le rouge et le proche infra-rouge. Tout particulièrement, le composé ou le nanocristal fluorescent peut émettre une fluorescence dans le visible et proche infra-rouge.In particular, the nano-crystals emit a fluorescence, which is dispersed over all fluorophores and is quenched or quenched simultaneously. By "simultaneously" is meant from 0.001 to 10 ns, in particular from 0.01 to 5 ns, in particular from 0.1 to 2 ns, or even from 0.5 to 1.5 ns. For the purpose of the present invention, the term "fluorescent" compound or nanocrystal means a compound or nanocrystal having the property of emitting, following a light excitation, electromagnetic radiation in the visible light domain or in the Near infrared (IR) - The excitation light signal can come from a light source such as a laser, an arc lamp or light-emitting diodes (LEDs). For example, the fluorescent compound or nanocrystal can emit electromagnetic radiation at wavelengths of 400 to 1200 nm, which corresponds to the window of relative transparency of living tissues. For example, the fluorescent compound or nanocrystal may emit electromagnetic radiation at wavelengths of 400 to 1000 nm, for example from 550 to 800 nm, which corresponds to fluorescence in the red and the near infra-red. In particular, the fluorescent compound or nanocrystal can emit fluorescence in the visible and near infra-red.
[0053] Les nano-cristaux peuvent être des nano-cristaux organiques ou organo-métalliques, en particulier être fluorescents, notamment comprendre au moins un, voire être constitués d'un, composé fluorescent à l'état cristallin. [0054] Les nano-cristaux peuvent comprendre au moins un composé molécule organique ou complexe de coordination qui fluoresce dans l'état solide et soluble dans un solvant organique.The nano-crystals may be organic or organometallic nano-crystals, in particular be fluorescent, in particular comprise at least one, or even consist of a fluorescent compound in the crystalline state. The nano-crystals may comprise at least one organic molecule compound or coordination complex which fluoresces in the solid state and is soluble in an organic solvent.
[0055] Selon l'invention, le composé organique ou organo-métallique fluorescent peut être par exemple choisi dans le groupe comprenant les composés fluorescents de la famille des polyaromatiques, par exemple le diphénylanthracène, le rubrène ou le tétracène, la famille des aromatiques, par exemple le 4-(N,N-diethylamino)-b- nitrostyrene (DEANST), 1 , 4-di(2 , 5-phenyloxazole) benzène (POPOP) ou le N-4-nitrophenyl-(L)-prolinol (NPP), la famille des stilbènes, par exemple le cyano-méthoxy-nitro- stilbène (CMONS), le diéthyl-amino-nitro-stilbène ou le stilbène, la famille des naphtilimides, par exemple le naphtilimide, la famille des rhodamines, par exemple la rhodamine B, la famille des auramines, par exemple l'auramine 0, la famille des pérylènes diimides, les dérivés du dipyrrométhène difluorure de bore, les complexes de terre rares, par exemple les complexes d'Europium.According to the invention, the fluorescent organic or organo-metallic compound may for example be chosen from the group comprising the fluorescent compounds of the polyaromatic family, for example diphenylanthracene, rubrene or tetracene, the aromatic family, for example 4- (N, N-diethylamino) -b-nitrostyrene (DEANST), 1,4-di (2,5-phenyloxazole) benzene (POPOP) or N-4-nitrophenyl- (L) -prolinol ( NPP), the stilbene family, for example cyano-methoxy-nitrosilbene (CMONS), diethyl-amino-nitro-stilbene or stilbene, the family of naphthimides, for example naphthimide, the family of rhodamines, for example Rhodamine B, the family of auramines, for example auramine O, the family of perylene diimides, derivatives of dipyrromethene difluoride boron, rare earth complexes, for example complexes Europium.
[0056] Le composé fluorescent à l'état solide, voire cristallin peut en particulier être choisi dans le groupe comprenant le cyanométhoxynitrostilbène, le naphtilimide, des molécules polyaromatiques, comme le rubrène ou le tétracène, les pérylènes diimides, le diphénylanthracène, les dérivés du dipyrrométhène difluorure de bore (Dipyrromethene Boron Difluoride - Bodipy®) , et les complexes d'Europium.The fluorescent compound in the solid state, or even crystalline, may in particular be selected from the group comprising cyanomethoxynitrostilbene, naphthimide, polyaromatic molecules, such as rubrene or tetracene, perylenes diimides, diphenylanthracene, derivatives of dipyrromethene boron difluoride (Dipyrromethene Boron Difluoride - Bodipy®), and Europium complexes.
[0057] Dans un mode de réalisation particulier, le composé fluorescent est choisi dans la famille des polyaromatiques, comme le rubrène, l'auramine 0 ou la rhodamine B, par exemple.In a particular embodiment, the fluorescent compound is selected from the polyaromatic family, such as rubrene, auramine O or rhodamine B, for example.
[0058] Les nano-cristaux peuvent être insérés dans les pores de la couche inorganique et/ou organo-minérale.The nano-crystals can be inserted into the pores of the inorganic and / or organo-mineral layer.
[0059] Par « pores », on entend au sens de la présente invention des interstices ou des vides entre les chaînes du réseau formant la couche inorganique et/ou organo-minérale (par exemple, le réseau silicate, titanate, zirconate, stannate, borate, aluminate, yttirate etc.) qui a polymerisé/polycondensé durant le procédé sol-gel.By "pores" is meant within the meaning of the present invention interstices or voids between the chains of the network forming the inorganic and / or organo-mineral layer (for example, the silicate network, titanate, zirconate, stannate, borate, aluminate, ytirate, etc.) which polymerized / polycondensed during the sol-gel process.
[0060] Les nano-cristaux observés par microscopie électronique en transmission peuvent être de forme sphérique et peuvent présenter un diamètre allant de 10 à 500 nm, notamment de 100 à 400 nm, en particulier de 150 à 300 nm.The nano-crystals observed by transmission electron microscopy may be spherical in shape and may have a diameter ranging from 10 to 500 nm, in particular from 100 to 400 nm, in particular from 150 to 300 nm.
[0061] Les nano-cristaux peuvent présenter une distribution en taille assez homogène, en particulier variant au maximum de +/- 20 %, voire +/- 10 %.The nano-crystals may have a relatively homogeneous size distribution, in particular varying at most by +/- 20%, or even +/- 10%.
[0062] Les nano-cristaux présents dans la couche inorganique et/ou organo-minérale présentent au moins une partie en contact direct avec l'extérieur, en particulier ils émergent de 2 à 50 % environ de leur diamètre par rapport à la couche inorganique et/ou organo-minérale, voire de 5 à 30 % de leur diamètre, en particulier lorsque les nanocristaux sont sphériques . La couche inorganique et/ou organo-minérale peut être une matrice sol-gel. [0063] Les nanocristaux peuvent dépasser de la couche inorganique et/ou organo-minérale de 1 à 100 nm, en particulier de 2 à 80 nm, voire de 3 à 50 nm. [0064] II suffit que les nanocristaux émergent un peu en surface pour être en contact direct avec l'extérieur (solution bio) soit de quelques nm à quelques dizaines de nm.The nano-crystals present in the inorganic and / or organo-mineral layer have at least a part in direct contact with the outside, in particular they emerge from 2 to 50% of their diameter relative to the inorganic layer. and / or organo-mineral, or even 5 to 30% of their diameter, in particular when the nanocrystals are spherical. The inorganic and / or organo-mineral layer may be a sol-gel matrix. The nanocrystals may exceed the inorganic and / or organo-mineral layer from 1 to 100 nm, in particular from 2 to 80 nm, or even from 3 to 50 nm. It suffices that the nanocrystals emerge a little surface to be in direct contact with the outside (organic solution) is a few nm to a few tens of nm.
[0065] Afin de présenter une résistance optimale, il est peut-être souhaitable de ne pas trop décaper la surface des couches sol-gel afin que les nanocristaux ne soient pas trop émergeants pour permettre une stabilisation mécanique excellente et ainsi éviter qu'ils se décrochent trop aisément de la couche sol-gel. Donc la hauteur d'émergence peut dépendre en partie du diamètre des nanocristaux. [0066] Ainsi, selon un mode de réalisation particulier, la hauteur d'émergence ne dépasse pas le tiers du diamètre des nanocristaux, notamment afin qu'ils demeurent bien ancrés dans la couche sol-gel. [0067] Cette mesure peut notamment être effectuée par microscopie à force atomique en mode oscillant intermittent en enregistrant la topographie de l'échantillon.In order to have optimum resistance, it may be desirable not to strip the surface of the sol-gel layers too much so that the nanocrystals are not too emerging to allow excellent mechanical stabilization and thus prevent them from drop too easily from the sol-gel layer. So the emergence height may depend in part on the diameter of the nanocrystals. Thus, according to a particular embodiment, the emergence height does not exceed one third of the diameter of the nanocrystals, especially so that they remain well anchored in the sol-gel layer. This measurement may in particular be performed by atomic force microscopy in intermittent oscillating mode by recording the topography of the sample.
[0068] Avantageusement, les nano-cristaux présents dans la couche inorganique et/ou organo-minérale sont solidement ancrés dans celle-ci, en particulier ils peuvent résister à une immersion dans une solution biologique à analyser (brins d'ADN, protéines...) et cela pendant quelques minutes. [0069] Par « résister », on entend que plus de 95 %, notamment plus de 99 %, voire plus de 99,9 %, des nanocristaux présents au début du test sont encore présents dans le (nano)matériau à la fin du test.Advantageously, the nano-crystals present in the inorganic and / or organo-mineral layer are firmly anchored in it, in particular they can withstand immersion in a biological solution to be analyzed (DNA strands, proteins. ..) and this for a few minutes. By "resist" means that more than 95%, especially more than 99% or more than 99.9% of the nanocrystals present at the beginning of the test are still present in the (nano) material at the end of the test.
[0070] Dans un mode de réalisation particulier, ces nano-cristaux peuvent présenter à leur surface au moins un type de sonde. Les nano-cristaux peuvent présenter un ou plusieurs types de sonde identiques ou différents, par exemple un, deux, trois, quatre ou cinq sondes identiques ou différentes, en particulier un, deux, trois, quatre ou cinq sondes de types différents. [0071] Au moins une sonde peut notamment être choisie dans le groupe comprenant les indicateurs colorés, l'ADN, les oligonucléotides ou polynucléotides, les polypeptides, les protéines, les anticorps, les sucres, les glycoprotéines et les lipides. Par exemple, les indicateurs colorés peuvent être sélectionnés parmi les indicateurs de pH, rédox et/ou complexation.In a particular embodiment, these nano-crystals may have at their surface at least one type of probe. Nano-crystals can have one or several types of identical or different probes, for example one, two, three, four or five identical or different probes, in particular one, two, three, four or five probes of different types. At least one probe may especially be chosen from the group comprising color indicators, DNA, oligonucleotides or polynucleotides, polypeptides, proteins, antibodies, sugars, glycoproteins and lipids. For example, the colored indicators can be selected from pH, redox and / or complexation indicators.
[0072] Cette sonde peut être simplement adsorbée ou bien greffée sur les nano-cristaux, en particulier via une liaison chimique. Par exemple, la sonde peut-être absorbée ou greffée via notamment une liaison covalente, ionique, électrostatique, ionocovalente, une liaison hydrogène, des interactions hydrophobes, les forces de van der waals. [0073] La sonde peut être chimiquement modifiée avec une fonction chimique adéquate de manière à pouvoir se greffer à la surface du nano-cristal.This probe can be simply adsorbed or grafted onto the nano-crystals, in particular via a chemical bond. For example, the probe can be absorbed or grafted via in particular a covalent, ionic, electrostatic, ionocovalent bond, a hydrogen bond, hydrophobic interactions, van der waals forces. The probe can be chemically modified with a suitable chemical function so as to be grafted on the surface of the nano-crystal.
[0074] Avantageusement, la sonde éventuellement chimiquement modifiée ne se lie pas à la surface de la couche sol-gel, notamment elle est insensible aux sites M- OH libres présents à la surface de la couche sol-gel. [0075] Ainsi, les nano-cristaux émergeants constituant, grâce à leur fluorescence, la fonction de signalisation, peuvent ainsi être fonctionnalisés par greffage d'une biomolécule portant une fonction sonde, tel demi-brin d'ADN, pour réaliser ainsi une bio-puce, capteur ou senseur comprenant un (nano)matériau selon l'invention, ou un support pour bio-puce, capteur ou senseur. Cet aspect de l'invention est discuté plus en détail dans les sections qui suivent. [0076] Selon un autre de ses aspects, l'invention a pour objet un procédé de fabrication de (nano)matériau selon l'invention, en particulier comprenant une couche inorganique et/ou organo-minérale, dans laquelle est intégré au moins un type de nano-cristal fluorescent organique ou organo-métallique émergeant de la surface de ladite couche de sorte qu'au moins une partie est en contact direct avec le milieu extérieur comprenant au moins l'étape suivante : a) dissolution d'une partie d'une couche inorganique et/ou organo-minérale de manière à faire émerger au moins une partie des nano-cristaux de la couche inorganique et/ou organo-minérale ou à augmenter la surface des nano-cristaux en contact avec le milieu extérieur. En particulier, dans l'étape de dissolution, ladite couche inorganique et/ou organo-minérale comprend ou inclut au moins un type de nano-cristal fluorescent.Advantageously, the optionally chemically modified probe does not bind to the surface of the sol-gel layer, in particular it is insensitive to free M-OH sites present on the surface of the sol-gel layer. Thus, emerging nano-crystals constituting, thanks to their fluorescence, the signaling function, can thus be functionalized by grafting a biomolecule carrying a probe function, such half-strand of DNA, to thereby achieve a bio chip, sensor or sensor comprising a (nano) material according to the invention, or a support for bio-chip, sensor or sensor. This aspect of the invention is discussed in more detail in the sections that follow. According to another of its aspects, the subject of the invention is a process for the manufacture of (nano) material according to the invention, in particular comprising an inorganic and / or organo-mineral layer, in which at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of said layer so that at least a portion is in direct contact with the external medium comprising at least the following step: a) dissolving a part of an inorganic and / or organo-mineral layer so as to emerge at least a portion of the nano-crystals of the inorganic and / or organo-mineral layer or to increase the surface of the nano-crystals in contact with the external medium. In particular, in the dissolution step, said inorganic and / or organo-mineral layer comprises or includes at least one type of fluorescent nano-crystal.
[0077] Cette étape de dissolution partielle de la couche sol-gel peut être effectuée par des agents, notamment chimiques, permettant de décaper de manière homogène la surface de la couche inorganique et/ou organo-minérale, en particulier sans détériorer les propriétés de fluorescence des nano-cristaux. [0078] Tout particulièrement, cette étape est effectuée par des agents permettant de décaper, choisis parmi des solutions aqueuses basiques, en particulier des bases inorganiques ou organiques, fortes ou faibles, notamment de soude, d'ammoniaque et/ou de potasse. Tout particulièrement ces solutions basiques présentent une concentration en base allant de 10"6 à 10"1 M, et notamment de 10"3 à 10"1 M.This step of partial dissolution of the sol-gel layer can be carried out by agents, in particular chemical agents, for homogeneously etching the surface of the inorganic and / or organo-mineral layer, in particular without deteriorating the properties of the fluorescence of nano-crystals. In particular, this step is performed by agents for pickling, selected from basic aqueous solutions, in particular inorganic or organic bases, strong or weak, including soda, ammonia and / or potassium hydroxide. Especially these basic solutions have a base concentration ranging from 10 "6-10" 1 million, including 10 "3-10" 1 M.
[0079] Ainsi, cette étape peut permettre un amincissement régulier de la couche inorganique et/ou organo-minérale, par décapage de la surface de la couche inorganique et/ou organo-minérale, et en particulier de matrices sol-gel silicatées.Thus, this step may allow a regular thinning of the inorganic and / or organo-mineral layer, by stripping the surface of the layer. inorganic and / or organo-mineral, and in particular silicate sol-gel matrices.
[0080] Tout particulièrement cette étape de dissolution contrôlée permet une vitesse d'amincissement allant de 5 à 100 nm/h, voire 10 à 50 nm/h.In particular this controlled dissolution step allows a thinning rate ranging from 5 to 100 nm / h, or 10 to 50 nm / h.
[0081] Le procédé selon l'invention peut également comprendre une étape consistant à préparer au moins une couche de matrice, en particulier de matrice sol-gel, comprenant ou incluant des nano-cristaux.The method according to the invention may also comprise a step of preparing at least one matrix layer, in particular of sol-gel matrix, comprising or including nano-crystals.
[0082] Ainsi, dans un mode de réalisation particulier, l'invention se rapporte à un procédé de fabrication de (nano)matériau comprenant une couche inorganique et/ou organo-minérale, dans laquelle est intégré au moins un type de nano-cristal fluorescent organique ou organo-métallique émergeant de la surface de ladite couche de sorte qu'au moins une partie est en contact direct avec le milieu extérieur, ledit procédé comprenant : a) une étape consistant à préparer au moins une couche inorganique et/ou organo-minérale comprenant ou incluant des nano-cristaux fluorescents organiques ou organo-métalliques, et b) une étape de dissolution d'une partie de ladite couche inorganique et/ou organo-minérale obtenue à l'étape a) de manière à faire émerger au moins une partie des nano-cristaux de la couche inorganique et/ou organo-minérale ou à augmenter la surface des nano- cristaux en contact avec le milieu extérieur.Thus, in a particular embodiment, the invention relates to a method of manufacturing (nano) material comprising an inorganic and / or organo-mineral layer, in which is integrated at least one type of nano-crystal organic or organometallic fluorescent emerging from the surface of said layer so that at least a portion is in direct contact with the external medium, said method comprising: a) a step of preparing at least one inorganic and / or organic layer -mineral comprising or including organic or organometallic fluorescent nano-crystals, and b) a step of dissolving a part of said inorganic and / or organo-mineral layer obtained in step a) so as to emerge at least part of the nano-crystals of the inorganic and / or organo-mineral layer or to increase the surface of the nano-crystals in contact with the external medium.
[0083] Dans un mode de réalisation, la couche inorganique et/ou organo-minérale est une couche sol-gel. Ainsi, l'étape a) peut comprendre une étape préalable (0) de préparation d'un mélange sol-gel comprenant : (i) une étape (Oa) de préparation d'un mélange sol-gel initial, par exemple en mélangeant dans un solvant au moins un composé organique ou organo-métallique fluorescent et au moins un alcoxyde métallique, par exemple de formule (I) , en présence d'eau ; et éventuellement ,In one embodiment, the inorganic and / or organo-mineral layer is a sol-gel layer. Thus, step a) may comprise a preliminary step (0) for preparing a sol-gel mixture comprising: (i) a step (Oa) for preparing an initial sol-gel mixture, for example by mixing in a solvent at least one fluorescent organic or organometallic compound and at least one metal alkoxide, for example of formula (I) in the presence of water; and eventually ,
(ii) une étape de stockage (Ob) du mélange sol-gel initial pendant une durée d afin de laisser réagir le mélange initial. [0084] Selon l'invention, la durée d peut être inférieure à plusieurs mois voire plusieurs années. Par exemple, la durée d peut être de 1 jour à 1 année, par exemple de 7 jours à 21 jours. En effet, le stockage du mélange permet de laisser vieillir le mélange afin de bien avancer les réactions d'hydrolyse et de condensation des alcoxydes métalliques .(ii) a storage step (Ob) of the initial sol-gel mixture for a period of time in order to allow the initial mixture to react. According to the invention, the duration d may be less than several months or even several years. For example, the duration d can be from 1 day to 1 year, for example from 7 days to 21 days. Indeed, the storage of the mixture allows the mixture to age to properly advance the hydrolysis and condensation reactions of metal alkoxides.
[0085] Selon l'invention, l'étape (0) de préparation du mélange sol-gel peut comprendre l'ajout d'un acide au mélange initial. En effet, l'acide, en abaissant le pH du mélange, permet de favoriser l'obtention de longues chaînes inorganiques qui sont favorables à la formation d'une couche inorganique et/ou organo-minérale dense autour des cristaux organiques ou organo-métalliques . Selon l'invention, le pH du mélange peut être de 1 à 7 , de préférence de 1 à 2.According to the invention, the step (0) for preparing the sol-gel mixture may comprise the addition of an acid to the initial mixture. Indeed, the acid, by lowering the pH of the mixture, makes it possible to promote the production of long inorganic chains which are favorable for the formation of a dense inorganic and / or organo-mineral layer around the organic or organometallic crystals. . According to the invention, the pH of the mixture can be from 1 to 7, preferably from 1 to 2.
[0086] On entend par « acide » au sens de la présente invention les acides de Brônsted, minéraux ou organiques. Parmi les acides utilisables, on peut citer par exemple l'acide chlorhydrique, l'acide nitrique ou l'acide acétique.For the purposes of the present invention, the term "acid" means the Brönsted acids, inorganic or organic. Among the acids that can be used, mention may be made, for example, of hydrochloric acid, nitric acid or acetic acid.
[0087] Le composé organique ou organo-métallique fluorescent, l' alcoxyde métallique et le solvant peuvent être choisis selon l'un quelconque des modes de réalisation décrits ci-dessus pour la mise en œuvre des nanomatériaux de l'invention.The fluorescent organic or organo-metallic compound, the metal alkoxide and the solvent may be selected according to any one of the embodiments described above for the implementation of the nanomaterials of the invention.
[0088] L'étape a) peut notamment être effectuée par la cristallisation de nano-cristaux organiques ou organo- métalliques fluorescents dans des couches minces de matrice sol-gel. Tout particulièrement cette étape peut être réalisée par centrifugation « à la tournette » ( « spin- coating » ) . [0089] Tout particulièrement, l'élaboration de cette couche inorganique et/ou organo-minérale, éventuellement sous forme de couche mince, peut être réalisée à température ambiante par hydrolyse et condensation d'une solution comprenant des précurseurs de la couche inorganique et/ou organo-minérale, notamment sol-gel, comme des alcoxydes de silicium, dans laquelle les molécules organiques ou organo-métalliques sont dissoutes dans un solvant, notamment organique. [0090] Par « organo-métallique », on entend des composés comprenant une partie organique et une partie métallique.Step a) may in particular be carried out by crystallization of organic nano-crystals or organometallic fluorescents in thin layers of sol-gel matrix. In particular, this step can be carried out by "spin-coating" centrifugation. In particular, the preparation of this inorganic and / or organo-mineral layer, optionally in the form of a thin layer, can be carried out at room temperature by hydrolysis and condensation of a solution comprising precursors of the inorganic layer and / or or organo-mineral, in particular sol-gel, such as silicon alkoxides, in which the organic or organometallic molecules are dissolved in a solvent, in particular an organic solvent. By "organo-metallic" is meant compounds comprising an organic part and a metal part.
[0091] La nano-cristallisation peut résulter d'une forte nucléation instantanée, suivie d'un contrôle de la croissance des nuclei par la viscosité et/ou la porosité du gel. La sursaturation de la phase organique peut être provoquée par l 'évaporâtion très rapide du solvant organique.The nano-crystallization can result from a strong instantaneous nucleation, followed by a control of the growth of nuclei by the viscosity and / or the porosity of the gel. The supersaturation of the organic phase can be caused by the very rapid evaporation of the organic solvent.
[0092] En particulier dans la procédure centrifugation à la tournette ( « spin-coating » ) , selon la vitesse de rotation, allant par exemple de 1000 à 4000 tr/mn, la viscosité du solvant et ou le temps de dépôt, des couches minces et/ou transparentes, par exemple de 100 à 1000 nm peuvent être réalisées.In particular in the spin spin coating procedure, depending on the rotational speed, for example from 1000 to 4000 rpm, the viscosity of the solvent and or the deposition time of the layers. Thin and / or transparent, for example 100 to 1000 nm can be made.
[0093] La couche inorganique et/ou organo-minérale de l'étape a) peut ensuite subir une étape de recuit, notamment à des températures voisines du point de fusion des nano-cristaux organiques. Cette étape de recuit est destinée à éliminer toute trace de solvant et stabiliser davantage la couche sol-gel, notamment inorganique (élimination de solvant résiduel, densification de la matrice). L'étape de recuit peut être conduite en particulier à des températures allant de 50 à 2500C, notamment de 80 à 1500C. [0094] Par « température voisine du point de fusion », on entend au sens de la présente invention une température comprise dans une gamme allant de point de fusion moins 5°C à point de fusion moins 1000C, en particulier de point de fusion moins 100C à point de fusion moins 500C. [0095] Cette étape de recuit peut permettre également d'améliorer la cristallinité des agrégats et/ou la stabilisation de la couche inorganique et/ou organo- minérale.The inorganic and / or organo-mineral layer of step a) can then undergo an annealing step, especially at temperatures close to the melting point of the organic nano-crystals. This annealing step is intended to eliminate any trace of solvent and further stabilize the sol-gel layer, in particular inorganic (residual solvent removal, densification of the matrix). The annealing step may be carried out especially at temperatures of from 50 to 250 0 C, in particular from 80 to 150 0 C. [0094] The term "temperature of the melting point" is meant within the meaning of the present invention a temperature ranging from a melting point of less than 5 ° C. to a melting point of less than 100 ° C., in particular of melting point less than 10 ° C. with a melting point of less than 50 ° C. [0095] This annealing step may also make it possible to improve the crystallinity of the aggregates and / or the stabilization of the inorganic and / or organo-mineral layer.
[0096] La taille des nano-cristaux peut ainsi être ajustée selon les conditions de nano-cristallisation confinée dans la couche inorganique et/ou organo-minérale, notamment dans la matrice sol-gel, en particulier silicatée.The size of the nano-crystals can thus be adjusted according to the conditions of nano-crystallization confined in the inorganic and / or organo-mineral layer, in particular in the sol-gel matrix, in particular silicate.
[0097] Dans la mesure où les nano-cristaux sont fluorescents, ils peuvent être visualisés grâce à des techniques comme la microscopie photonique confocale. Ceci est illustré par la figure 2 qui est une image par microscopie photonique confocale de nano-cristaux de rubrène en matrice sol-gel.As far as the nano-crystals are fluorescent, they can be visualized by techniques such as confocal photonic microscopy. This is illustrated in FIG. 2 which is an image by confocal photon microscopy of nano-rubrene crystals in sol-gel matrix.
[0098] Dans le cas où les nano-cristaux présentent une taille inférieure à 100 nm, la microscopie électronique en transmission peut être utilisée pour observer la distribution et la taille des nano-cristaux. Ceci est illustré par la Figure 3 qui est une image par microscopie électronique en transmission de nano-cristaux de rubrène en matrice sol-gel.In the case where the nano-crystals have a size smaller than 100 nm, transmission electron microscopy can be used to observe the distribution and the size of the nano-crystals. this is illustrated by Figure 3 which is an image by transmission electron microscopy of rubrene nano-crystals in sol-gel matrix.
[0099] Ces visualisations directes des nano-cristaux peuvent permettre d'ajuster les paramètres influant sur la cristallisation confinée des chromophores (molécules organiques ou organo-métalliques fluorescentes), en particulier sur la nature des précurseurs permettant de former la matrice sol-gel, par exemple la nature des alcoxydes de silicium, les cinétiques d'hydrolyse et de condensation, la nature et les taux de solvants, la concentration en fluorophores, la sursaturation et les paramètres de centrifugation.These direct visualizations of the nano-crystals can make it possible to adjust the parameters influencing the confined crystallization of the chromophores (organic or organo-metallic fluorescent molecules), in particular on the nature of the precursors making it possible to form the sol-gel matrix, for example the nature of the silicon alkoxides, the kinetics of hydrolysis and condensation, the nature and levels of solvents, the concentration of fluorophores, the supersaturation and the centrifugation parameters.
[0100] Selon un autre de ses objets, la présente invention concerne un film de (nano)matériau selon l'invention. Dans un mode de réalisation particulier, il s'agit d'un film comprenant une matrice sol-gel comprenant des nano-cristaux stabilisés mécaniquement, lesdits nano- cristaux émergeant de la surface dudit film de sorte qu'au moins une partie de chaque nano-cristal est en contact direct avec le milieu extérieur. En particulier le (nano)matériau est une matrice sol-gel comprenant des nano- cristaux stabilisés mécaniquement, lesdits nano-cristaux émergeant de la surface dudit film de sorte qu'au moins une partie de chaque nano-cristal est en contact direct avec le milieu extérieur, disposé sur un support solide, de type lame de verre par exemple. [0101] Les techniques de préparation de tels films sont connues de l'homme du métier, et ne seront pas développées dans la présente demande. Dans ce contexte, plusieurs méthodes de dépôt de couches minces sol-gel peuvent être utilisées : techniques de centrifugation du substrat (plusieurs milliers de tours par minute). On peut également utiliser d'autres techniques telles que le tirage du substrat verticalement (« dip-coating » en anglais). Le lecteur pourra se référer à l'ouvrage de Brinker et Scherer précité [Ref I].According to another of its objects, the present invention relates to a film of (nano) material according to the invention. In a particular embodiment, it is a film comprising a sol-gel matrix comprising mechanically stabilized nano-crystals, said nano-crystals emerging from the surface of said film so that at least a portion of each nano -crystal is in direct contact with the external environment. In particular, the (nano) material is a sol-gel matrix comprising mechanically stabilized nano-crystals, said nano-crystals emerging from the surface of said film so that at least a portion of each nano-crystal is in direct contact with the nano-crystal. external medium, arranged on a solid support, such as glass slide for example. The techniques for preparing such films are known to those skilled in the art, and will not be developed in the present application. In this context, several methods of deposition of thin sol-gel layers can be used: substrate centrifugation techniques (several thousand revolutions per minute). It is also possible to use other techniques such as drawing the substrate vertically ("dip-coating" in English). The reader will be able to refer to the work of Brinker and Scherer mentioned above [Ref I].
[0102] Comme discuté auparavant, les dispositifs basés sur la détection d'un signal fluorescent sont notamment utilisés dans les domaines reposant sur la détection de réactions chimiques ou biologiques (par exemple les biopuces) .As discussed above, devices based on the detection of a fluorescent signal are used in particular in the areas based on the detection of chemical or biological reactions (for example biochips).
[0103] Les biopuces sont un outil biochimique de collection massive d'information notamment sur les acides nucléiques (puces ADN) et les acides aminés (puces à protéines), les antigènes et anticorps (capteurs immunologiques ) . Associées à des techniques de traitement numérique des informations récoltées, les puces ADN permettent de mener les recherches (détection, séparation, identification, étude) permettant d'accéder directement aux ADN. [0104] Les puces à protéines permettent de détecter, identifier, séparer, étudier les protéines et déterminer leurs activités, fonctions, interactions, modifications au cours du temps . [0105] Les capteurs immunologiques basés sur une liaison avec une enzyme permettent de détecter des anticorps/antigènes .Biochips are a biochemical tool for mass collection of information including nucleic acids (DNA chips) and amino acids (protein chips), antigens and antibodies (immunological sensors). Combined with digital processing techniques harvested information, DNA chips can conduct research (detection, separation, identification, study) to access directly to the DNA. The protein chips can detect, identify, separate, study the proteins and determine their activities, functions, interactions, changes over time. The immunological sensors based on binding with an enzyme make it possible to detect antibodies / antigens.
[0106] D'une manière générale, un domaine d'application de l'invention concerne donc les puces, notamment les biopuces, capteurs et senseurs, qui se présentent sous forme d'un support solide à la surface duquel des éléments biochimiques sont immobilisés.In general, a field of application of the invention therefore relates to chips, including biochips, sensors and sensors, which are in the form of a solid support on the surface of which biochemical elements are immobilized. .
[0107] Ainsi, la présente invention trouve notamment des applications dans la fabrication de supports pour les biopuces capteurs ou senseurs. Il s'agit notamment de supports biopuces compatibles avec le greffage d'acides nucléiques (biopuces à ADN, biopuces à ARN), d'acides aminés (puces à protéines, puces immunologiques ) , ainsi que les biopuces cellulaires utilisées notamment dans les études de transfection.Thus, the present invention finds particular applications in the manufacture of media for sensor or sensor biochips. These include microarray supports compatible with the grafting of nucleic acids (DNA microarrays, RNA biochips), amino acids (protein chips, immunological chips), as well as the cellular biochips used in particular in transfection studies.
[0108] Ainsi, selon un de ses aspects, l'invention concerne un support pour biopuce, capteur ou senseurs comprenant un substrat dont une surface comprend une couche de (nano)matériau ou de film selon l'invention. Dans un mode de réalisation, le (nano)matériau comprend au moins une couche inorganique et/ou organo-minérale, dans laquelle est intégré au moins un type de nano-cristal fluorescent organique ou organo-métallique émergeant de la surface de ladite couche de sorte qu'au moins une partie du nano- cristal est en contact direct avec le milieu extérieur.Thus, according to one of its aspects, the invention relates to a support for a biochip, sensor or sensor comprising a substrate whose surface comprises a layer of (nano) material or film according to the invention. In one embodiment, the (nano) material comprises at least one inorganic and / or organo-mineral layer, in which at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of said so that at least a part of the nano-crystal is in direct contact with the external medium.
[0109] Dans un mode de réalisation, le support comprend un substrat sur lequel le (nano)matériau de la présente invention est déposé, notamment sous forme de couche mince ou de film. Il peut s'agir de tout matériau solide connu de l'homme du métier, tel que par exemple les matériaux supports utilisés pour la fabrication des microsystèmes d'analyse et des biopuces. Il peut s'agir d'un substrat organique ou inorganique. Il peut être constitué par exemple d'un matériau choisi dans le groupe comprenant du verre, de la silice, du polycarbonate, du nymon, du polyméthylméthacrylate (PMMA), du polystyrène, du copolymère cyclooléfine (COC), et du polystyrène acrylonitrile (SAN). Le support est typiquement une lame de verre de taille comparable à une lame de microscope.In one embodiment, the support comprises a substrate on which the (nano) material of the present invention is deposited, in particular in the form of a thin layer or film. It may be any solid material known to those skilled in the art, such as, for example, the support materials used for the manufacture of analytical microsystems and biochips. It can be an organic or inorganic substrate. It may consist for example of a material selected from the group consisting of glass, silica, polycarbonate, nymon, polymethyl methacrylate (PMMA), polystyrene, cycloolefin copolymer (COC), and acrylonitrile polystyrene (SAN). ). The support is typically a glass slide of comparable size to a microscope slide.
[0110] Le substrat peut être préalablement nettoyé afin d'améliorer l'adhérence de la couche de (nano)matériau à sa surface. Ce nettoyage peut être par exemple un nettoyage chimique, éventuellement suivi d'un traitement thermique. Les techniques de nettoyage en particulier celles utilisées pour la préparation de supports de biopuce, capteur et senseur, connues de l'homme du métier sont utilisables. Pour ce nettoyage, on peut utiliser tout solvant approprié pour dépoussiérer et/ou dégraisser la surface du substrat, de préférence sans l'abîmer. On peut citer par exemple comme solvant de nettoyage le trichloréthylène, l'acétone, l'alcool éthylique, l'eau désionisée, des solutions acides ou basiques, etc. Le nettoyage peut consister à baigner le substrat dans un ou plusieurs de ces solvants successivement. Le nettoyage est alors généralement suivi d'un séchage. Le nettoyage peut également consister tout simplement à débarrasser le substrat de ses poussières par un jet d'air comprimé.The substrate may be previously cleaned in order to improve the adhesion of the (nano) material layer to its surface. This cleaning can be for example a cleaning chemical, possibly followed by a heat treatment. Cleaning techniques, in particular those used for the preparation of biochip carriers, sensor and sensor, known to those skilled in the art are usable. For this cleaning, any solvent suitable for dusting and / or degreasing the surface of the substrate may be used, preferably without damaging it. Trichlorethylene, acetone, ethyl alcohol, deionized water, acidic or basic solutions, etc. may be mentioned as cleaning solvents, for example. The cleaning may consist of bathing the substrate in one or more of these solvents successively. Cleaning is usually followed by drying. The cleaning may also simply consist of ridding the substrate of its dust with a jet of compressed air.
[0111] La couche de (nano)matériau peut être déposée sur la surface du substrat par toute technique connue de 1 ' homme du métier pour déposer ce type de matériau sur une surface. Tout particulièrement, lorsque le (nano)matériau comprend une couche sol-gel, cette étape peut être réalisée par :The (nano) material layer may be deposited on the surface of the substrate by any technique known to those skilled in the art for depositing this type of material on a surface. In particular, when the (nano) material comprises a sol-gel layer, this step can be carried out by:
(i) une étape (Oa) de préparation d'un mélange initial, par exemple en mélangeant dans un solvant au moins un composé organique ou organo-métallique fluorescent et au moins un alcoxyde métallique, par exemple de formule (I) , en présence d'eau ; éventuellement ,(i) a step (Oa) for preparing an initial mixture, for example by mixing in a solvent at least one fluorescent organic or organometallic compound and at least one metal alkoxide, for example of formula (I), in the presence water; eventually ,
(ii) une étape de stockage (Ob) du mélange initial pendant une durée d afin de laisser réagir le mélange initial, et(ii) a step of storing (Ob) the initial mixture for a period of time so as to allow the initial mixture to react, and
(iii) une étape de dépôt du mélange sol-gel obtenu à l'étape (ii) sur un support, et (iv) une étape de centrifugation « à la tournette » ( « spin-coating » ) pour obtenir une couche inorganique et/ou organo-minérale comprenant des nano-cristaux organique sou organo-métalliques fluorescents à la surface du support.(iii) a step of depositing the sol-gel mixture obtained in step (ii) on a support, and (iv) a "spin-coating" centrifugation step to obtain an inorganic and / or organo-mineral layer comprising fluorescent organic organometallic organic nano-crystals on the surface of the support.
[0112] En effet, un mélange sol-gel étant préparé par hydrolyse et condensation de sels ou d'alcoxydes de métaux ou de métalloides M, avantageusement en présence d'un solvant organique, ce solvant organique peut faciliter le dépôt et le séchage de la couche de (nano)matériau sur le substrat.Indeed, a sol-gel mixture being prepared by hydrolysis and condensation of salts or alkoxides of metals or metalloids M, preferably in the presence of an organic solvent, this organic solvent can facilitate the deposition and drying of the layer of (nano) material on the substrate.
[0113] Dans un mode de réalisation, la couche de (nano)matériau selon l'invention peut être déposée sur ladite surface du substrat à une épaisseur de 50 nm à 1000 nm, voire 100 nm à 1000 nm. En fait, on comprend aisément que 1 ' épaisseur minimale est une épaisseur qui permet avantageusement de couvrir la surface du substrat, sans laisser de trous. Quant à l'épaisseur maximale, il est de façon générale préférable de ne pas aller au-delà du micromètre. En effet, si l'épaisseur est supérieure au micromètre, des problèmes de craquage/fissuration de la couche liés à l 'évaporation du solvant peuvent survenir (tensions internes dues aux forces capillaires ; voir ouvrage de référence de Brinker et Scherer [ref I]) . il est toutefois superflu d'utiliser des quantités trop importantes de (nano)matériau pour mettre en oeuvre la présente invention. [0114] Une fois la couche inorganique et/ou organo- minérale comprenant des nano-cristaux organiques ou organo- métalliques fluorescents déposés sur le support, celui-ci peut être en outre soumis à une étape de dissolution d'une partie d'une couche inorganique et/ou organo-minérale de manière à faire émerger au moins une partie des nano- cristaux de la couche inorganique et/ou organo-minérale ou à augmenter la surface des nano-cristaux en contact avec le milieu extérieur.In one embodiment, the (nano) material layer according to the invention may be deposited on said surface of the substrate at a thickness of 50 nm to 1000 nm, or even 100 nm to 1000 nm. In fact, it is easy to understand that the minimum thickness is a thickness which advantageously makes it possible to cover the surface of the substrate without leaving holes. As for the maximum thickness, it is generally preferable not to go beyond the micrometer. In fact, if the thickness is greater than one micrometer, problems of cracking / cracking of the layer related to the evaporation of the solvent can occur (internal tensions due to capillary forces, see reference book of Brinker and Scherer [ref I] ). however, it is unnecessary to use too large amounts of (nano) material to carry out the present invention. Once the inorganic and / or organo-mineral layer comprises fluorescent organic or organometallic nanocrystals deposited on the support, the latter may be furthermore subjected to a step of dissolving part of a inorganic and / or organo-mineral layer so as to bring out at least a portion of the nanoparticles crystals of the inorganic and / or organo-mineral layer or to increase the surface of the nano-crystals in contact with the external medium.
[0115] Les divers modes de réalisation décrits ci-dessus pour la préparation du (nano)matériau selon l'invention sont applicables à la mise en œuvre des supports pour puces, capteurs ou senseurs (e.g., solvants, composés fluorescents, alcoxydes métalliques, etc.). [0116] En particulier, un ou plusieurs types de sonde identiques ou différents peuvent être greffés ou adsorbées sur la partie des nano-cristaux émergeant de la surface de la couche inorganique et/ou organo-minérale. Pour ce faire, on pourra appliquer les divers modes de réalisation décrits précédemment pour le (nano)matériau de l'invention (i.e., types de sonde, modes de liaison, etc.).The various embodiments described above for the preparation of the (nano) material according to the invention are applicable to the implementation of supports for chips, sensors or sensors (eg, solvents, fluorescent compounds, metal alkoxides, etc.). In particular, one or more types of identical or different probe may be grafted or adsorbed on the part of the nano-crystals emerging from the surface of the inorganic and / or organo-mineral layer. To do this, we can apply the various embodiments described above for the (nano) material of the invention (i.e., types of probe, modes of connection, etc.).
[0117] Dans un mode de réalisation particulier, le support selon l'invention peut comprendre un empilement de couches minces diélectriques formant un miroir de Bragg intercalé entre le substrat et la couche de nanomatériau. Pour la mise en œuvre de cet aspect de l'invention, le lecteur pourra se référer aux documents brevet de la société Genewave, par exemple WO 2004/042376 [Ref 5] et WO 2007/045755 [Ref 6]. Il s'agit de substrats présentant en surface de réseau un miroir de Bragg pour aire en sorte que le signal fluorescent soit complètement réfléchi vers le détecteur pour augmenter la sensibilité du système. Ces substrats sont communément appelés « amplislides » . [0118] Cette configuration particulière peut permettre d'augmenter le champ excitateur à l'intérieur de la couche de nanomatériau. Ainsi, cette configuration a pour effet d'augmenter l'excitation des nano-cristaux organiques ou organo-métalliques luminescents ancrés dans la couche de nanomatériau, et donc d'augmenter la quantité de lumière émise dans le substrat. On précise qu'un miroir de Bragg est un empilement successif de plusieurs couches minces diélectriques d'indices de réfraction n: et n2 différents. L'épaisseur de chacune de ces couches est égale à λ/(4n), avec n pouvant prendre la valeur n: ou n2. La variable 1 correspond à la longueur d'onde à laquelle on souhaite avoir une réflexion maximale pour le miroir de Bragg. [0119] Selon un autre de ses aspects, l'invention se rapporte à l'utilisation du (nano)matériau selon l'invention pour la fabrication de supports pour puces, notamment des supports pour biopuces, capteurs ou senseurs.In a particular embodiment, the support according to the invention may comprise a stack of thin dielectric layers forming a Bragg mirror interposed between the substrate and the nanomaterial layer. For the implementation of this aspect of the invention, the reader can refer to the patent documents of Genewave, for example WO 2004/042376 [Ref 5] and WO 2007/045755 [Ref 6]. These are substrates having a Bragg mirror at the grating surface so that the fluorescent signal is completely reflected back to the detector to increase the sensitivity of the system. These substrates are commonly called "amplislides". This particular configuration may make it possible to increase the exciter field inside the nanomaterial layer. Thus, this configuration has the effect of increasing the excitation of luminescent organic or organometallic nano-crystals anchored in the nanomaterial layer, and therefore of increasing the quantity of light emitted into the substrate. We specify that a Bragg mirror is a successive stack of several thin dielectric layers of refractive indices n : and n 2 different. The thickness of each of these layers is equal to λ / (4n), with n being able to take the value n : or n 2 . The variable 1 corresponds to the wavelength at which it is desired to have a maximum reflection for the Bragg mirror. According to another of its aspects, the invention relates to the use of the (nano) material according to the invention for the manufacture of media for chips, including supports for biochips, sensors or sensors.
[0120] Selon encore un autre de ses aspects, l'invention se rapporte à une puce, notamment une bio-puce, un capteur ou un senseur comprenant au moins un (nano)matériau ou un film selon l'invention. Les puces, notamment les biopuces, capteurs et senseurs et leurs divers modes de mise en oeuvre sont connus de l'homme de métier, ont fait l'objet de nombreux rapports et publications. À titre illustratif, on pourra se référer à (i) D. L. Gerhold, « Better Therapeutics through microarrays, Nature Genetics, 32, p 547-552 (2002) [Ref 7] ; (ii) M.A. Shipp « Diffuse large B- cell lymphoma outcome prédiction by gene-expression profiling and supervised machine learning » Nature Médecine 8 pp 68-74 (2002) [Ref 8] ; and (iii) L. Quijada, M. Soto and J. M. Requena « Genomic DNA microarrays as a tool of gène expression in leishmania » Expression Parasitology, 111, pp 64-70 (2005) [Ref 9].According to yet another of its aspects, the invention relates to a chip, in particular a bio-chip, a sensor or a sensor comprising at least one (nano) material or a film according to the invention. Chips, including biochips, sensors and sensors and their various modes of implementation are known to those skilled in the art, have been the subject of numerous reports and publications. For illustrative purposes, reference can be made to (i) D. L. Gerhold, "Better Therapeutics through Microarrays, Nature Genetics, 32, p 547-552 (2002) [Ref 7]; (ii) M.A. Shipp "Broad Diffuse B-cell Lymphoma Outcome Prediction by Gene-Expression Profiling and Supervised Machine Learning" Nature Medicine 8 pp 68-74 (2002) [Ref 8]; and (iii) L. Quijada, M. Soto and J. M. Requena "Genomic DNA microarrays as a tool of expression gene in leishmania" Expression Parasitology, 111, pp 64-70 (2005) [Ref 9].
[0121] S ' agissant d'une puce ADN par exemple, on peut greffer ou adsorber un ou plusieurs oligonucléotides ou polynucléotides sur les nano-cristaux organiques ou organo- métalliques luminescents émergeant de la surface de la couche inorganique et/ou organo-minérale du (nano)matériau selon l'invention. Avantageusement, ces oligonucléotides ou polynucléotides sont des oligonucléotides ou polynucléotides simples brins spécifiques connus. Leur rôle est de détecter des cibles marquées complémentaires, présentes dans le mélange complexe à analyser. En effet, le principe de détection utilisé dans les puces ADN repose sur les possibilités d' appariement de brins d'ADN avec leurs bases complémentaires. Ces oligonucléotides ou polynucléotides sont autant de sondes qui permettent d'hybrider des séquences d'ADN complémentaires provenant d'un échantillon biologique à analyser. Avantageusement, les nano-cristaux organiques ou organo-métalliques luminescents sont fluorescents. Une fois la puce hybridée, la détection des chaînes est effectuée par mesure du signal fluorescent correspondant. Les images obtenues sont numérisées pour être ensuite traitées par des algorithmes de traitement spécifiques de ces données, mis en oeuvre par des moyens informatiques. [0122] Les divers modes de réalisation décrits ci-dessus pour la préparation du (nano)matériau et supports pour puces, capteurs et senseurs selon l'invention sont applicables à la mise en œuvre des puces, capteurs ou senseurs (e.g., solvants, composés fluorescents, alcoxydes métalliques, etc.). [0123] En particulier, un ou plusieurs types de sonde identiques ou différents peuvent être greffés ou adsorbés sur la partie des nano-cristaux émergeant de la surface de la couche inorganique et/ou organo-minérale. Pour ce faire, on pourra appliquer les divers modes de réalisation décrits précédemment pour le (nano)matériau de l'invention (i.e., types de sonde, modes de liaison, etc...)-In the case of a DNA chip, for example, one or more oligonucleotides or polynucleotides can be grafted onto or adsorbed onto the luminescent organic or organometallic nanocrystals emerging from the surface of the inorganic and / or organo-mineral layer. (nano) material according to the invention. Advantageously, these oligonucleotides or polynucleotides are known single-stranded oligonucleotides or specific polynucleotides. Their role is to detect complementary labeled targets present in the complex mixture to be analyzed. Indeed, the detection principle used in DNA chips is based on the possibility of pairing DNA strands with their complementary bases. These oligonucleotides or polynucleotides are all probes that make it possible to hybridize complementary DNA sequences originating from a biological sample to be analyzed. Advantageously, the organic nano-crystals or organo-metallic luminescent are fluorescent. Once the hybridized chip, the chain detection is performed by measuring the corresponding fluorescent signal. The images obtained are digitized and then processed by specific processing algorithms of these data, implemented by computer means. The various embodiments described above for the preparation of the (nano) material and supports for chips, sensors and sensors according to the invention are applicable to the implementation of chips, sensors or sensors (eg, solvents, fluorescent compounds, metal alkoxides, etc.). In particular, one or more types of identical or different probes may be grafted or adsorbed on the part of the nano-crystals emerging from the surface of the inorganic and / or organo-mineral layer. To do this, it will be possible to apply the various embodiments described above for the (nano) material of the invention (ie, types of probe, modes of connection, etc.).
[0124] Le (nano)matériau selon l'invention peut permettre d'obtenir une meilleure sensibilité (capacité de détection), et par exemple permettre de gagner jusqu'à 2 aThe (nano) material according to the invention can make it possible to obtain a better sensitivity (capacity of detection), and for example to make it possible to win up to 2
3 ordres de grandeur par rapport aux biopuces actuelles. [0125] Ce gain en sensibilité peut notamment être dû à la conjugaison de deux facteurs principaux : [0002] chaque nano-cristal peut être composé d'un très grand nombre de fluorophores (de 104 à 1010 fluorophores par nano-cristal selon son diamètre), et3 orders of magnitude compared to current biochips. This gain in sensitivity may notably be due to the combination of two main factors: [0002] each nano-crystal may be composed of a very large number of fluorophores (from 10 4 to 10 10 fluorophores per nano-crystal according to its diameter), and
[0003] au transfert d'énergie donneur-donneur entre fluorophores, ainsi l'excitation lumineuse peut explorer un large domaine sur une distance de 8 nm en moyenne, soit deux fois le rayon de Fôrster, distance de transfert d'énergie entre un donneur (fluorophore isolé) et un accepteur d'énergie.At the donor-donor energy transfer between fluorophores, and the light excitation can explore a wide range over a distance of 8 nm on average, twice the radius of Fôrster, energy transfer distance between a donor (isolated fluorophore) and an energy acceptor.
[0126] Ainsi, la sensibilité peut être améliorée d'un facteur 100 ou plus par rapport à un fluorophore isolé. [0127] Cette amélioration peut être constatée en mesurant le nombre de mole de fluorophores éteint dans une suspension aqueuse de nanocristaux par mole d'inhibiteur de fluorescence. L'effet d'inhibition observé (modification du déclin de fluorescence) est beaucoup plus important dans le cas des nano-cristaux.Thus, the sensitivity can be improved by a factor of 100 or more compared to an isolated fluorophore. This improvement can be observed by measuring the number of moles of fluorophores extinguished in an aqueous suspension of nanocrystals per mole of fluorescence inhibitor. The observed inhibition effect (modification of fluorescence decline) is much greater in the case of nano-crystals.
[0128] Tout particulièrement les puces à fluorescence selon l'invention peuvent présenter une capacité de détection (sensibilité) d'une molécule cible pour cent molécules fluorescentes. Cette capacité de détection, ou sensibilité, se mesure mieux au moyen de cristaux en solution où les concentrations sont mieux connues. Un exemple de ce mode de réalisation est donné dans l'Exemple[0128] In particular, the fluorescence chips according to the invention may have a detection capacity (sensitivity) of a target molecule for one hundred fluorescent molecules. This detection capacity, or sensitivity, is better measured by means of crystals in solution where the concentrations are better known. An example of this embodiment is given in the Example
4 ci-dessous.4 below.
Brève description des figures [0129] La figure 1 représente un exemple de tracé de « Stern Volmer » de Io/I dont la pente donne le nombre de molécules de rubrène dans la nanoparticule éteintes par chaque molécule de quencheur, le bleu de cibacron (CB) par exemple. Autrement dit, il s'agit du nombre moyen de molécules de rubrène par nano-cristal organique fluorescent qui sont éteintes par chaque molécule de quencheur. [0130] La figure 2 représente un exemple d'image par microscopie photonique confocale de nano-cristaux de rubrène en matrice sol-gel issue de précurseur TMOS/TMSE.Brief description of the figures FIG. 1 represents an example of a "Stern Volmer" plot of Io / I whose slope gives the number of rubrene molecules in the nanoparticle extinguished by each quencher molecule, for example, cibacron blue (CB). In other words, it is the average number of rubrene molecules per fluorescent organic nano-crystal that are extinguished by each quencher molecule. FIG. 2 represents an example of an image by confocal photon microscopy of rubrene nano-crystals in sol-gel matrix derived from TMOS / TMSE precursor.
[0131] La figure 3 représente un exemple d'image par microscopie électronique en transmission de nano-cristaux de rubrène en matrice sol-gel issue de précurseur TMOS/TMSE [0132] La figure 4 représente un exemple d'images enregistrées au microscope à force atomique (AFM) en mode intermittent montrant des nanocristaux de rubrène émergeant d'environ 35 nm de la surface d'une couche mince sol-gel issue de précurseur TMOS/TMSE. [0133] La figure 5 représente un exemple d'images enregistrées au microscope à force atomique en champ proche (AFM) montrant des nanocristaux de rhodamine B émergeant de 2 à 3 nm de la surface d'une couche mince sol-gel silicatée issue de précurseur TMOS/GPTMS. [0134] La figure 6 représente un exemple d'images enregistrées au microscope à force atomique en champ proche (AFM) montrant des nanocristaux d'auramine 0 émergeant de 15 à 20 nm de la surface d'une couche mince sol-gel silicatée issue de précurseur TMOS/TMSE.FIG. 3 represents an exemplary image by transmission electron microscopy of rubrene nano-crystals in a sol-gel matrix derived from TMOS / TMSE precursor. FIG. 4 represents an example of images recorded under a microscope at atomic force (AFM) in intermittent mode showing rubrene nanocrystals emerging about 35 nm from the surface of a sol-gel thin layer derived from TMOS / TMSE precursor. FIG. 5 represents an example of images recorded using a near-field atomic force microscope (AFM) showing nanocrystals of rhodamine B emerging from 2 to 3 nm from the surface of a silicate sol-gel thin layer originating from TMOS / GPTMS precursor. FIG. 6 represents an example of images recorded using a near-field atomic force microscope (AFM) showing auramine 0 nanocrystals emerging from 15 to 20 nm from the surface of a silicate sol-gel thin layer resulting from TMOS / TMSE precursor.
EXEMPLESEXAMPLES
[0135] Le procédé de dissolution partielle de la surface des couches sol-gel pour rendre émergeant les nanocristaux organiques selon l'invention a été appliqué sur différentes couches nanocomposites afin d'illustrer l'aspect générique de l'invention. Des décapages ont été mis en œuvre sur trois couches sol-gel différentes contenant des nanocristaux organiques différents. Ceux-ci sont détaillés dans les Exemples 1 à 3 ci-dessous.The method of partially dissolving the surface of the sol-gel layers to make the organic nanocrystals emerging according to the invention was applied to different nanocomposite layers in order to illustrate the generic aspect. of the invention. Stripping was carried out on three different sol-gel layers containing different organic nanocrystals. These are detailed in Examples 1 to 3 below.
[0136] Exemple 1. NANOCRISTAUX DE RUBRENE EN COUCHES MINCES SOL-GEL SILICATEES ISSUES DE PRECURSEUR TMOS + TMSE [0137] La poudre microcristalline de rubrène (1,8 mg ; 0,033 mmoles) a été dissoute dans une solution contenant 4,570 mL de tétrahydrofurane, 0,257 mL de tétraméthylsiloxane (TMOS), 0,222 mL de 1,2- bis(triméthoxysilyl)ethane (TMSE) et 0,219 mL d'une solution HCl 0,1 M. Les proportions molaires dans le mélange initial sont : 2 TMOS + 1 TMSE + 48 THF + 10 H2O + 0.005 rubrène. (H2O se réfère à la quantité d'eau introduite avec la solution HCl 0,1 M).EXAMPLE 1 SILICATE THIN-LAYER THERMO-GEL RUBRENE NANOCRISTALS FORMED FROM PRECURSOR TMOS + TMSE The microcrystalline rubrene powder (1.8 mg, 0.033 mmol) was dissolved in a solution containing 4.570 mL of tetrahydrofuran 0.257 mL of tetramethylsiloxane (TMOS), 0.222 mL of 1,2-bis (trimethoxysilyl) ethane (TMSE) and 0.219 mL of 0.1 M HCl solution. The molar proportions in the initial mixture are: 2 TMOS + 1 TMSE + 48 THF + 10H 2 O + 0.005 rubrene. (H 2 O refers to the amount of water introduced with 0.1M HCl solution).
[0138] La solution a ensuite été stockée pendant plusieurs jours afin de bien avancer les réactions d'hydrolyse et de condensation des alcoxydes. On note que le mélange sol-gel est très stable. On peut donc éventuellement laisser le mélange en stockage pendant plusieurs semaines, voire plusieurs mois si on le souhaite. Cependant, plusieurs jours sont suffisants pour bien faire hydrolyse et condensation des alcoxydes de départ.The solution was then stored for several days in order to properly advance the hydrolysis and condensation reactions of the alkoxides. It is noted that the sol-gel mixture is very stable. It may therefore be possible to leave the mixture in storage for several weeks, or even several months if desired. However, several days are sufficient to properly hydrolyze and condensation of starting alkoxides.
[0139] Un volume de 200μL du mélange ainsi obtenu est déposé sur une lame de verre de microscope. [0140] La lame de microscope est ensuite introduite dans un Spin coater de la société Suss Microtech de type RC8 GYRSET. La couche mince sol-gel est obtenue par spin- coating en utilisant les conditions suivantes : [0141] vitesse de rotation : 4000 tr/min [0142] accélération : 3000 tr/min2 [0143] durée de la rotation : 10sA volume of 200 .mu.l of the mixture thus obtained is deposited on a slide of microscope glass. The microscope slide is then introduced into a spin coater of the company Suss Microtech type RC8 GYRSET. The sol-gel thin layer is obtained by spin-coating using the following conditions: [0141] rotational speed: 4000 rpm [0142] acceleration: 3000 rpm 2 [0143] duration of the rotation: 10s
[0144] La couche mince ainsi obtenue est ensuite recuite à 1000C entre 24h et 72h. [0145] On obtient ainsi des nanocristaux de rubrène de 200 nm de diamètre moyen inclus dans une couche mince de 250 nm d'épaisseur. Les nanocristaux ont été visualisés par microscopie optique confocale (figure 2) et microscopie électronique en transmission (figure 3).The thin layer thus obtained is then annealed at 100 ° C. between 24 and 72 hours. We thus obtain rubrene nanocrystals of 200 nm in mean diameter included in a thin layer of 250 nm in thickness. The nanocrystals were visualized by confocal optical microscopy (FIG. 2) and transmission electron microscopy (FIG. 3).
[0146] La couche mince nanocomposites incrustées de nanocristaux de rubrène est ensuite soumise à des conditions de dissolutions contrôlées. Ainsi, la couche mince est décapée pour faire apparaître les nanocristaux de rubrène en surface en la trempant dans une solution de NaOH diluée (10"3M) durant 16H.The nanocomposite thin layer encrusted with rubrene nanocrystals is then subjected to controlled dissolution conditions. Thus, the thin layer is etched to expose the surface rubrene nanocrystals by soaking in a dilute NaOH solution (10 -3 M) for 16 hours.
[0147] Les nanocristaux émergeant à la surface de la couche sol-gel sont présentés en figure 4. Cette figure permet de visualiser directement une lame supportant une couche sol-gel de très faible rugosité (0,5 nm RMS) qui supportent des nanocristaux émergeants qui sont la fonction de signalisation de ce nouveau type de détecteur. [0148] Les images de la figure 4 ont été enregistrées au microscope à force atomique (AFM) en mode intermittent montrant des nanocristaux de rubrène émergeant à la surface de la couche mince sol-gel. Ces images montrent que les nanocristaux sont bien en contact avec le milieu extérieur mais qu'ils restent bien ancrés dans la couche. Il sont décapés à moins de 50% de leur diamètre, et de l'ordre de 30% au maximum. Les nanocristaux émergent d'environ 35 nm au-dessus de la couche sol-gel. On n'observe pas de perte (trou) de nanocristaux sur ces couches décapées qui sont ainsi prêtes à être fonctionnalisées. [0149] Ces échantillons constituent un exemple de réalisation de base (fonction de signalisation de ce nouveau type de biopuce). Ainsi, nous avons pu maîtriser par dissolution lente (2-3 nm/h) le décapage de la surface de nos couches sol-gel permettant l'émergence des nanocristaux organiques tout en conservant la même rugosité d'environ 0.5 nm en RMS au niveau de la couche silicatée. Cette conservation de la très faible rugosité de la couche sol-gel illustre bien le parfait contrôle du décapage qui est très homogène en épaisseur sur toute la couche selon un procédé de dissolution par monocouches moléculaire, ce qui est tout à fait remarquable.The nanocrystals emerging on the surface of the sol-gel layer are shown in FIG. 4. This figure makes it possible to directly view a slide supporting a sol-gel layer of very low roughness (0.5 nm RMS) that supports nanocrystals. emerging which are the signaling function of this new type of detector. The images of FIG. 4 were recorded using an atomic force microscope (AFM) in intermittent mode showing rubrene nanocrystals emerging on the surface of the sol-gel thin layer. These images show that the nanocrystals are in good contact with the external environment but that they remain well anchored in the layer. They are stripped to less than 50% of their diameter, and of the order of 30% maximum. The nanocrystals emerge about 35 nm above the sol-gel layer. There is no loss (hole) of nanocrystals on these stripped layers which are ready to be functionalized. These samples are an example of a basic embodiment (signaling function of this new type of biochip). Thus, we were able to control by slow dissolution (2-3 nm / h) the etching of the surface of our sol-gel layers allowing the emergence of organic nanocrystals while maintaining the same roughness of about 0.5 nm in RMS at silicate layer. This preservation of the very low roughness of the sol-gel layer illustrates the perfect control of the etching which is very homogeneous in thickness over the entire layer according to a method of dissolution by molecular monolayers, which is quite remarkable.
[0150] Exemple 2. NANOCRISTAUX DE RHODAMINE B EN COUCHES MINCES SOL-GEL SILICATEES ISSUES DE PRECURSEUR TMOS/GPTMS [0151] Un mélange sol-gel a été préparé selon un mode opératoire similaire à celui de l'Exemple 1, en utilisant 21,7 mg de poudre microcristalline de rhodamine B 3,662 mL de MeOH, 0,267 mL de tétraméthylsiloxane (TMOS), 0,599 mL de 3-glycidoxypropyl)trimethoxysilane (GPTMS) et 0,277 mL d'une solution HCl 0,1 M. Les proportions molaires dans le mélange initial sont : 0.4 TMOS + 0.6 GPTMS + 20 MeOH + 8 H2O + 0.01 rhodamine B. (H2O se réfère à la quantité d'eau introduite avec la solution HCl 0,1 M). [0152] Le mélange ainsi obtenu a été stocké pendant plusieurs jours afin de bien avancer les réactions d'hydrolyse et de condensation des alcoxydes. On note que le mélange sol-gel est très stable. On peut donc éventuellement laisser le mélange en stockage pendant plusieurs semaines, voire plusieurs mois si on le souhaite. Cependant, plusieurs jours sont suffisants pour bien faire hydrolyse et condensation des alcoxydes de départ. [0153] Une couche mince sol-gel a été réalisée sur lame de microscope par spin-coating dans des conditions de spin coating et de recuit identiques à celles de l'Exemple 1. [0154] On obtient ainsi des nanocristaux de rhodamine B de 40 nm de diamètre moyen inclus dans une couche mince silicatée de 180 nm d'épaisseur.EXAMPLE 2 RHODAMINE B NANOCRYSTALS IN THIN LAYERS GEL SILICATE FROM PRECURSOR TMOS / GPTMS [0151] A sol-gel mixture was prepared according to a procedure similar to that of Example 1, using 21 7 mg of microcrystalline rhodamine B powder, 3.662 mL of MeOH, 0.267 mL of tetramethylsiloxane (TMOS), 0.599 mL of 3-glycidoxypropyl) trimethoxysilane (GPTMS) and 0.277 mL of 0.1M HCl solution. The molar proportions in the initial mixture are: 0.4 TMOS + 0.6 GPTMS + 20 MeOH + 8 H 2 O + 0.01 rhodamine B. (H 2 O refers to the amount of water introduced with 0.1M HCl solution). The mixture thus obtained was stored for several days to advance the reactions of hydrolysis and condensation of the alkoxides. It is noted that the sol-gel mixture is very stable. It may therefore be possible to leave the mixture in storage for several weeks, or even several months if desired. However, several days are sufficient to properly hydrolyze and condensation of starting alkoxides. A sol-gel thin layer was produced on a microscope slide by spin-coating under spin coating and annealing conditions identical to those of Example 1. [0154] Thus, rhodamine B nanocrystals were obtained. 40 nm in average diameter included in a silicate thin layer 180 nm thick.
[0155] La lame est ensuite plongée dans une solution de NaOH à 1.10~3mol/L pendant 4h. [0156] Les nanocristaux émergent, après décapage de 2 à 3 nm (voir Figure 5).The blade is then immersed in a solution of NaOH at 1.10 ~ 3 mol / L for 4 hours. The nanocrystals emerge after etching of 2 to 3 nm (see FIG. 5).
[0157] La vitesse de dissolution pour cette matrice et avec NaOH à 1.10~3mol/L est d'environ 2 nm par heure confirmant les résultats de l'exemple précédent.The dissolution rate for this matrix and with NaOH at 1.10 ~ 3 mol / L is about 2 nm per hour confirming the results of the previous example.
[0158] Exemple 3. NANOCRISTAUX D 'AURAMINE O EN COUCHES MINCES SOL-GEL SILICATEES ISSUES DE PRECURSEUR TMOS/TMSE [0159] Un mélange sol-gel a été préparé selon un mode opératoire similaire à celui de l'Exemple 1, en utilisant 42,36 mg de poudre microcristalline d'auramine 0 3,528 mL de THF, 0,257 mL de tétraméthylsiloxane (TMOS), 0,222 mL de l,2-bis(triméthoxysilyl)ethane (TMSE) et 0,219 mL d'une solution HCl 0,1 M. Les proportions molaires dans le mélange initial sont : 2 TMOS + 1 TMSE + 75 THF + 10 H2O + 0.04 auramine 0. (H2O se réfère à la quantité d'eau introduite avec la solution HCl 0,1 M).EXAMPLE 3 AURAMINE O NANO-CRYSTALS IN SILICATED THIN-GEL THIN LAYERS FROM PRECURSOR TMOS / TMSE A sol-gel mixture was prepared according to a procedure similar to that of Example 1, using 42.36 mg of auramine microcrystalline powder 0 3.528 mL of THF, 0.257 mL of tetramethylsiloxane (TMOS), 0.222 mL of 1,2-bis (trimethoxysilyl) ethane (TMSE) and 0.219 mL of a 0.1 HCl solution M. The molar proportions in the initial mixture are: 2 TMOS + 1 TMSE + 75 THF + 10 H 2 O + 0.04 auramine 0. (H 2 O refers to the amount of water introduced with 0.1M HCl solution ).
[0160] Le mélange ainsi obtenu a été stocké pendant plusieurs jours ( 3 ou 4 jours) afin de bien avancer les réactions d'hydrolyse et de condensation des alcoxydes. Comme dans les exemples précédents, le mélange sol-gel est stable. On peut donc éventuellement laisser le mélange en stockage pendant plusieurs semaines, voire plusieurs mois si on le souhaite. Cependant, plusieurs jours sont suffisants pour bien faire hydrolyse et condensation des alcoxydes de départ.The mixture thus obtained was stored for several days (3 or 4 days) to advance the reactions of hydrolysis and condensation of the alkoxides. As in the previous examples, the sol-gel mixture is stable. It may therefore be possible to leave the mixture in storage for several weeks, or even several months if desired. However, several days are sufficient to properly hydrolyze and condense the starting alkoxides.
[0161] Une couche mince sol-gel a été réalisée sur lame de microscope par spin-coating dans des conditions de spin coating et de recuit identiques à celles des exemples précédents .A sol-gel thin layer was made on a microscope slide by spin-coating under spin coating and annealing conditions identical to those of the previous examples.
[0162] On obtient ainsi des nanocristaux d'auramine 0 de 200 à 300 nm de diamètre moyen inclus dans une couche mince silicatée de 300 nm d'épaisseur.Auramine 0 nanocrystals of 200 to 300 nm in mean diameter are thus obtained, which are included in a silicate thin film 300 nm thick.
[0163] La lame est ensuite plongée dans une solution de NaOH à 1.10"1ITiOlZL pendant 1H30. On note que la méthode de décapage fonctionne également avec une solution nettement plus concentrée que celles utilisées dans les exemples précédents. La vitesse de dissolution s'en trouve plus élevée.The slide is then immersed in a solution of NaOH at 1.10 -1 ITiOlZL for 1H30 It is noted that the etching method also works with a solution much more concentrated than those used in the preceding examples. found higher.
[0164] Après décapage, Les nanocristaux émergent de 15 à 20 nm (voir Figure 6). La vitesse de dissolution pour une telle matrice et pour [NaOH] = 1.10"1ITiOlZL est d'environ 8- 10 nm par heure.After stripping, the nanocrystals emerge from 15 to 20 nm (see FIG. 6). The dissolution rate for such a matrix and for [NaOH] = 1.10 "1 ITiOlZL is about 8-10 nm per hour.
[0165] Les trois exemples précédents démontrent l'applicabilité de l'invention à diverses couches sol-gel, et différents types de nano-cristaux, de tailles différentesThe three preceding examples demonstrate the applicability of the invention to various sol-gel layers, and different types of nano-crystals, of different sizes.
Exemple 4Example 4
[0166] Une suspension aqueuse de nanocristaux est préparée. 10 mL d'une solution de rubrène à 1,3 mmolZL dans un mélange l 'éthanolZTHF 3 :7 est injectée avec une seringue avec une petite aiguille dans 100 mL d'une solution aqueuse de CTACl (chlorure de cethyl trimethyl ammonium) à 5 mmolZL. La solution est filtrée à travers un filtre millipore de taille coupure de 50 nm. La concentration en molécule fluorescente est mesurée par absorption dans une cuvette en quartz de 1 cm de trajet optique. La fluorescence est mesurée dans un spectrofluorimètre SPEX Fluorolog avec une excitation à 495 nm avec des fentes de 2 nm à l'émission et à l'excitation. On mesure l'intensité de fluorescence pour des ajouts connus de « quencheur » (molécule sonde qui inhibe la fluorescence des nanocristaux par transfert d'énergie non radiatif entre le nano-cristal initialement excité et cette molécule se trouvant en surface du nano-cristal). Un quencheur typique est le bleu de cibachron. Un autre est le bleu de méthylène. Tous deux sont dissous à 1 mmol/L dans l'eau. On montre sur la figure 1 qu'avec un quencheur pour 100 molécules de rubrène on a déjà 33% d'extinction de la fluorescence : Le tracé de "Stern Volmer" de Io/I est une droite en accord avec une adsorption du quencheur, le bleu de cibacron (CB), sur les nanocristaux de rubrène dans cet exemple. La pente donne le nombre de molécule de rubrène dans la nanoparticule dont la fluorescence est éteinte par chaque molécule de quencheur. [0166] An aqueous suspension of nanocrystals is prepared. 10 ml of a solution of rubrene at 1.3 mmolZL in a mixture of ethanol ZTHF 3: 7 is injected with a syringe with a small needle into 100 ml of an aqueous solution of CTACl (cethyltrimethylammonium chloride) at 5 ml. mmolZL. The solution is filtered through a millipore filter of cut size of 50 nm. The concentration of fluorescent molecule is measured by absorption in a quartz cuvette of 1 cm optical path. The fluorescence is measured in a SPEX Fluorolog spectrofluorometer with excitation at 495 nm with 2 nm slots at emission and excitation. The fluorescence intensity is measured for known "quencher" additions (probe molecule which inhibits the fluorescence of the nanocrystals by non-radiative energy transfer between the initially excited nano-crystal and this molecule lying on the surface of the nano-crystal) . A typical quencher is cibachron blue. Another is methylene blue. Both are dissolved at 1 mmol / L in water. It is shown in FIG. 1 that with a quencher for 100 molecules of rubrene, there is already 33% extinction of the fluorescence: the "Stern Volmer" plot of Io / I is a straight line in agreement with a adsorption of the quencher, the blue of cibacron (CB), on the nanocrystals of rubrene in this example. The slope gives the number of rubrene molecules in the nanoparticle whose fluorescence is extinguished by each quencher molecule.
Liste des référencesList of references
(1) CJ. Brinker et G.W. Scherer, Sol-Gel Science, The Physics and Chemistry of SoIGeI Processing, Académie Press, New York, 1990.(1) CJ. Brinker and G.W. Scherer, Sol-Gel Science, The Physics and Chemistry of Processing, Academy Press, New York, 1990.
(2) D. Avnir, V.R. Kaufman and R. Reisfeld, J. Non. Cryst. Solids, 1985, 74, 395-406.(2) D. Avnir, V.R. Kaufman and R. Reisfeld, J. No. Cryst. Solids, 1985, 74, 395-406.
(3) C. Sanchez and F. Ribot, New J. Chem. , 1994, 18, 1007- 1047. (4) Brevet français FR 2 853 307.(3) C. Sanchez and F. Ribot, New J. Chem. , 1994, 18, 1007-1047. (4) French patent FR 2 853 307.
(5) WO 2004/042376.(5) WO 2004/042376.
(6) WO 2007/045755.(6) WO 2007/045755.
(7) D. L. Gerhold, « Better Therapeutics through microarrays, Nature Genetics, 32, p 547-552 (2002). (8) M.A. Shipp « Diffuse large B-cell lymphoma outeome prédiction by gene-expression profiling and supervised machine learning » Nature Médecine 8 pp 68-74 (2002).(7) D. L. Gerhold, "Better Therapeutics Through Microarrays, Nature Genetics, 32, p 547-552 (2002). (8) M.A. Shipp "Diffuse Wide B-cell Lymphoma Uteome Prediction by Gene-Expression Profiling and Supervised Machine Learning" Nature Medicine 8 pp 68-74 (2002).
(9) L. Quijada, M. Soto and J. M. Requena « Genomic DNA microarrays as a tool of gène expression in leishmania » Expression Parasitology, 111, pp 64-70 (2005) .(9) L. Quijada, M. Soto and J. M. Requena "Genomic DNA microarrays as a gene expression tool in leishmania" Expression Parasitology, 111, pp 64-70 (2005).
Chacune des références ci-dessus est incorporée par la présente, par référence, dans son intégralité. Each of the above references is hereby incorporated by reference in its entirety.

Claims

REVENDICATIONS
1. Matériau comprenant au moins une couche inorganique et/ou organo-minérale, dans laquelle est intégré au moins un type de nano-cristal fluorescent organique ou organo- métallique émergeant de la surface de ladite couche de sorte qu'au moins une partie du nano-cristal est en contact direct avec le milieu extérieur.1. Material comprising at least one inorganic and / or organo-mineral layer, in which at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of said layer is integrated so that at least a part of the nano-crystal is in direct contact with the external environment.
2. Matériau selon la revendication 1 dans lequel la couche inorganique et/ou organo-minérale est une matrice sol-gel de variété Silicate, Titanate, Zirconate, Stannate, Borate, Aluminate ou Yttriate.2. Material according to claim 1 wherein the inorganic and / or organo-mineral layer is a sol-gel matrix of Silicate, Titanate, Zirconate, Stannate, Borate, Aluminate or Yttriate variety.
3. Matériau selon la revendication 1 ou 2 se présentant sous forme de film.3. Material according to claim 1 or 2 in the form of a film.
4. Matériau selon l'une quelconque des revendications 1 à 3 dans lequel ledit au moins nano-cristal comprend au moins une molécule organique ou un complexe de coordination qui fluoresce dans l'état solide et qui est soluble dans un solvant organique, la molécule organique ou complexe de coordination étant choisie dans le groupe comprenant les composés fluorescents de la famille des polyaromatiques, la famille des aromatiques, la famille des stilbènes, la famille des naphtilimides, la famille des rhodamines, la famille des auramines, la famille des pérylènes diimides, les dérivés du dipyrrométhène difluorure de bore, et les complexes de terre rares.4. Material according to any one of claims 1 to 3 wherein said at least nano-crystal comprises at least one organic molecule or coordination complex which fluoresces in the solid state and which is soluble in an organic solvent, the molecule organic or coordination complex being chosen from the group comprising the fluorescent compounds of the polyaromatic family, the aromatic family, the stilbene family, the family of naphthimides, the family of rhodamines, the family of auramines, the family of perylenes diimides , dipyrromethene boron difluoride derivatives, and rare earth complexes.
5. Matériau selon la revendication 4 dans lequel la molécule organique ou complexe de coordination est choisie dans le groupe comprenant le diphénylanthracène, le rubrène, le tétracène, le cyano-méthoxy-nitro-stilbène (CMONS), le diéthyl-amino-nitro-stilbène, le naphtilimide, la rhodamine B, l'auramine 0, et les complexes d'Europium.5. Material according to claim 4 wherein the organic molecule or coordination complex is selected from the group comprising diphenylanthracene, rubrene, tetracene, cyano-methoxy-nitro-stilbene (CMONS), diethyl-amino-nitro-stilbene, naphthimide, rhodamine B, auramine O, and Europium complexes.
6. Matériau selon l'une quelconque des revendications 1 à 5 dans lequel ledit au moins nano-cristal présente à sa surface au moins un type de sonde.6. Material according to any one of claims 1 to 5 wherein said at least nano-crystal has on its surface at least one type of probe.
7. Matériau selon la revendication 6 dans lequel au moins une sonde est choisie dans le groupe comprenant les indicateurs colorés, l'ADN, les oligonucléotides ou polynucléotides, les polypeptides, les protéines, les anticorps, les sucres, les glycoprotéines et les lipides.The material of claim 6 wherein at least one probe is selected from the group consisting of color indicators, DNA, oligonucleotides or polynucleotides, polypeptides, proteins, antibodies, sugars, glycoproteins and lipids.
8. Matériau selon la revendication 6 ou 7 dans lequel au moins une sonde est greffée sur ledit au moins nano- cristal .8. The material of claim 6 or 7 wherein at least one probe is grafted onto said at least nano-crystal.
9. Matériau selon l'une quelconque des revendications 1 à 8 dans lequel ledit au moins nano-cristal présente un diamètre allant de 10 à 500 nm.9. Material according to any one of claims 1 to 8 wherein said at least nano-crystal has a diameter ranging from 10 to 500 nm.
10. Matériau selon l'une quelconque des revendications 1 à 9 dans lequel ledit au moins un type de nano-cristal présente une distribution en taille variant au maximum de +/- 20 %.10. Material according to any one of claims 1 to 9 wherein said at least one type of nano-crystal has a size distribution of up to +/- 20%.
11. Matériau selon l'une quelconque des revendications 1 à 10 dans lequel ledit au moins un type de nano-cristal émerge de 2 à 50 % environ de son diamètre par rapport à la couche inorganique et/ou organo-minérale.11. Material according to any one of claims 1 to 10 wherein said at least one type of nano-crystal emerges from 2 to 50% of its diameter relative to the inorganic and / or organo-mineral layer.
12. Film de matériau caractérisé en ce que le matériau est tel que défini selon l'une quelconque des revendications 1 à 11. 12. Film material characterized in that the material is as defined in any one of claims 1 to 11.
13. Film selon la revendication 12 disposé sur un support solide.13. The film of claim 12 disposed on a solid support.
14. Film selon la revendication 12 ou 13 ayant une épaisseur allant de 50 à 1500 nm.The film of claim 12 or 13 having a thickness of from 50 to 1500 nm.
15. Procédé de fabrication de matériau comprenant une couche inorganique et/ou organo-minérale, dans laquelle est intégré au moins un type de nano-cristal fluorescent organique ou organo-métallique émergeant de la surface de ladite couche de sorte qu'au moins une partie est en contact direct avec le milieu extérieur, ledit procédé comprenant : a) une étape consistant à préparer au moins une couche inorganique et/ou organo-minérale comprenant ou incluant des nano-cristaux fluorescents organiques ou organo- métalliques, et b) une étape de dissolution d'une partie de ladite couche inorganique et/ou organo-minérale obtenue à l'étape a) de manière à faire émerger au moins une partie des nano- cristaux de la couche inorganique et/ou organo-minérale ou à augmenter la surface des nano-cristaux en contact avec le milieu extérieur.A process for producing a material comprising an inorganic and / or organo-mineral layer, in which at least one type of organic or organometallic fluorescent nano-crystal emerging from the surface of said layer is integrated so that at least one part is in direct contact with the external environment, said process comprising: a) a step of preparing at least one inorganic and / or organo-mineral layer comprising or including organic or organometallic fluorescent nano-crystals, and b) a step of dissolving a portion of said inorganic and / or organo-mineral layer obtained in step a) so as to bring out at least a portion of the nano-crystals of the inorganic and / or organo-mineral layer or to increase the surface of the nano-crystals in contact with the external environment.
16. Procédé selon la revendication 15 dans lequel l'étape de dissolution est effectuée par des agents permettant de décaper de manière homogène la surface de la couche inorganique et/ou organo-minérale.16. The method of claim 15 wherein the dissolving step is performed by agents for homogeneously etching the surface of the inorganic and / or organo-mineral layer.
17. Procédé selon la revendication 16 dans lequel les agents permettant de décaper sont des solutions aqueuses basiques fortes ou faibles. 17. The method of claim 16 wherein the stripping agents are strong or weak basic aqueous solutions.
18. Procédé selon l'une quelconque des revendications 15 à 17 dans lequel l'étape de dissolution est contrôlée pour permettre une vitesse d'amincissement de la couche inorganique et/ou organo-minérale, par décapage de la surface de la couche inorganique et/ou organo-minérale, allant de 5 à 100 nm/h.18. A method according to any one of claims 15 to 17 wherein the dissolving step is controlled to allow a rate of thinning of the inorganic and / or organo-mineral layer, by stripping the surface of the inorganic layer and or organo-mineral, ranging from 5 to 100 nm / h.
19. Procédé selon l'une quelconque des revendications 15 à 18 dans lequel la couche inorganique et/ou organo- minérale de l'étape a) est obtenue par la cristallisation de nano-cristaux organiques ou organo-métalliques fluorescents dans des couches minces de matrice sol-gel.19. Process according to claim 15, in which the inorganic and / or organo-mineral layer of step a) is obtained by crystallization of fluorescent organic or organometallic nano-crystals in thin films of sol-gel matrix.
20. Procédé selon la revendication 19 dans lequel la matrice obtenue subit une étape de recuit.20. The method of claim 19 wherein the matrix obtained undergoes an annealing step.
21. Support pour biopuce, capteur et/ou senseur comprenant un substrat dont une surface comprend une couche de matériau tel que défini selon l'une quelconque des revendications 1 à 11 ou un film tel que défini selon la revendication 12, 13 ou 14.21. Support for a biochip, sensor and / or sensor comprising a substrate whose surface comprises a layer of material as defined according to any one of claims 1 to 11 or a film as defined according to claim 12, 13 or 14.
22. Puce, capteur ou senseur comprenant au moins un matériau tel que défini selon l'une quelconque des revendications 1 à 11 ou un film tel que défini selon la revendication 12, 13 ou 14.22. A chip, sensor or sensor comprising at least one material as defined in any one of claims 1 to 11 or a film as defined in claim 12, 13 or 14.
23. Puce, capteur ou senseur selon la revendication 22 présentant une capacité de détection d'une molécule cible pour cent molécules fluorescentes. 23. A chip, sensor or sensor according to claim 22 having a detection capacity of a target molecule per 100 fluorescent molecules.
PCT/FR2008/050490 2007-03-21 2008-03-21 Fluorescent organic nanocrystals for producing biosensors WO2008145875A1 (en)

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EP08805659A EP2125992B1 (en) 2007-03-21 2008-03-21 Fluorescent organic nanocrystals for producing biosensors
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9346997B2 (en) 2008-05-08 2016-05-24 Board Of Regents Of The University Of Texas System Luminescent nanostructured materials for use in electrogenerated chemiluminescence

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103765215A (en) * 2011-06-07 2014-04-30 生命科技公司 Fluorogenic semiconductor nanocrystals
WO2013033049A1 (en) * 2011-08-26 2013-03-07 Aviana Molecular Technologies, Llc Biocoated piezoelectric biosensor platform for point-of-care diagnostic use
JP6188711B2 (en) * 2011-12-01 2017-08-30 ザ ユニバーシティ オブ ユタ リサーチ ファウンデイション Photonic element on flat and curved substrate and method of manufacturing the same
US9429769B2 (en) * 2013-05-09 2016-08-30 Johnson & Johnson Vision Care, Inc. Ophthalmic device with thin film nanocrystal integrated circuits
DK178111B1 (en) * 2013-10-24 2015-05-26 Københavns Uni Sol-gel based matrix

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6770220B1 (en) * 1999-07-15 2004-08-03 Presens Precision Sensing Gmbh Production and use of luminescent microparticles and nanoparticles
EP1541656A1 (en) * 2002-06-19 2005-06-15 National Institute of Advanced Industrial Science and Technology Semiconductor superfine particle phosphor and light emitting device

Family Cites Families (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
FR2846745B1 (en) 2002-10-30 2004-12-24 Genewave DEVICE FOR SUPPORTING CHROMOPHORE ELEMENTS.
FR2853307B1 (en) 2003-04-07 2006-07-07 Centre Nat Rech Scient PROCESS FOR PREPARING A COMPOSITION OF NANOPARTICLES OF AT LEAST ONE CRYSTALLINE METAL OXIDE
FR2854696A1 (en) 2003-05-06 2004-11-12 Commissariat Energie Atomique BIOPUCE SUPPORT USING THIN LAYERS OF SOL GEL MATERIAL AND METHOD OF MAKING SAME
FR2892196B1 (en) 2005-10-18 2008-06-20 Genewave Soc Par Actions Simpl METHOD FOR MANUFACTURING INTEGRATED DETECTION BIOSENSOR

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6770220B1 (en) * 1999-07-15 2004-08-03 Presens Precision Sensing Gmbh Production and use of luminescent microparticles and nanoparticles
EP1541656A1 (en) * 2002-06-19 2005-06-15 National Institute of Advanced Industrial Science and Technology Semiconductor superfine particle phosphor and light emitting device

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
BOTZUNG-APPERT ET AL: "Spatial control of organic nanocrystal nucleation in sol-gel thin films for 3-D optical data storage devices or chemical multi-sensors", JOURNAL OF CRYSTAL GROWTH, ELSEVIER, AMSTERDAM, NL, vol. 283, no. 3-4, 1 October 2005 (2005-10-01), pages 444 - 449, XP005065765, ISSN: 0022-0248 *
HE H ET AL: "Synthesis and room temperature photoluminescence of AgI nanoparticles embedded in silica sol-gel coating", SOLID STATE IONICS, NORTH HOLLAND PUB. COMPANY. AMSTERDAM, NL, vol. 175, no. 1-4, 30 November 2004 (2004-11-30), pages 651 - 654, XP004667657, ISSN: 0167-2738 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9346997B2 (en) 2008-05-08 2016-05-24 Board Of Regents Of The University Of Texas System Luminescent nanostructured materials for use in electrogenerated chemiluminescence

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